Yufei Hua's research while affiliated with Jiangnan University and other places
The water absorption of soybeans during soaking is directly related to the quality characteristics and the flavor properties of soybeans for processing. In this paper, the effects of water absorption of soybean seed on the quality of soymilk and the release of flavor compounds were investigated during soaking at 4 °C, 25 °C, and 50 °C at different pH values. The results showed that the water absorption rate increased as the soaking temperature and pH increased, while the equilibrium value was relatively stable. Peleg's equation with good fitting of the absorption kinetics was used to predict the hydration characteristics of undehulled soybean. MALDI-TOF/TOF-MS results showed that the major released proteins are basic 7S globulin, which is released in large amounts at high temperature. The water absorption of soybean seed significantly enhanced the extraction yields of protein, fat and solids of the prepared soymilk, and alkaline soaking pH further promoted the extraction of proteins and solids. A high soaking temperature can significantly decrease the required soaking time; however, it is unfavorable to the extraction yields of fat, proteins and solids, as well as the whiteness values and the particle sizes. The beany odor compounds of soymilk mainly consisted of hexanal, trans-2-hexenal, 1-octene-3-ol, hexanol, and 2-pentylfuran, and their contents were positively correlated with soaking temperature. A good balance of soymilk quality and flavor compound release can be achieved with soaking conditions of 25 °C and pH 9.
Hydrophobic interactions and disulfide bonds involved in heat‐induced soy protein gels were characterised by determining the dissolution kinetics of gels. Reducing SDS‐PAGE results revealed that all proteins in gel network could be dissolved simultaneously by 1% (w/v) SDS solution, while a majority of glycinin (11S) A polypeptide and a moderate amount of 11S‐B polypeptides, 7S‐α′, α, γ, and β subunits were found in 2% (w/v) DTT dissolving samples. Stronger interaction force between proteins in gel network would result in lower dissolution constant rate. The breaking force of soy gels increased from 543 to 2171 gforce with increasing heating temperature from 85 to 100 °C, and denaturation of 11S globulins played an important role in the development of gel network. As increasing heating time from 30 to 120 min, the breaking force of gels increased from 1687 to 2175 gforce, then decreased to 1253 gforce when the time was prolonged to 240 min. Negative correlations were observed between breaking force and dissolution constant rate kSDS or Δk, which suggested that the strengthening of both hydrophobic interactions and disulfide bonds.
Purpose: The purpose of this paper is to evaluate the changes in descriptive sensory properties and overall consumer acceptability of soymilk prepared from roasted soybeans. Design/methodology/approach: In total, 12 purposively selected post graduate students majoring in Food Science conducted descriptive sensory analysis after being trained for 18 h in sensory analysis, while 75 untrained students conducted consumer acceptability test of soymilk prepared by roasting soybeans at a temperature of 110°C for 20, 40, 60, 80 and 100 min and at 120°C for 20 min. Findings: Results have revealed that roasting soybeans improved sensory properties by significantly (p<0.05) decreasing the objectionable green, beany flavours and increasing sweet taste, viscosity and roasted flavour. Furthermore, results from the principal component analysis revealed that aroma and sweet taste were the most critical sensory attributes. In addition, it was found out that soymilk samples prepared by roasting soybeans at 110°C for 40 and 60 min and at 120°C for 20 min were significantly more acceptable than the control soymilk. Research limitations/implications: The participants in this study were from one locality and predominantly soybean consuming community and therefore there is need to conduct the study in a different locality in order to validate the study findings. Practical implications: The study can assist small scale processors that might not have access to lipoxygenase-free soybeans and other technologies for improving the quality of soymilk. Social implications: The study can be used as a guide for connecting the food processers with the external world of consumption. Originality/value: For the first time, the study findings have demonstrated that controlled soybean roasting can be a useful strategy for improving soymilk sensory properties and consumer acceptability. The findings in this study can be usefully used in the quality control of soy bean-based products.
In this study, to recover proteins and minimize anti-nutritional factor (ANF) levels in soybean wastewater, a novel quick-shearing system was used to obtain homogeneous electrostatic complexes between soybean whey protein (SWP) and a high concentration (4%, w/v) of polysaccharide. During processing, protein recovery and trypsin inhibitory activity loss induced by complexing with sulfated and carboxylated polysaccharides were investigated as function of polysaccharide type, pH, protein to polysaccharide mass ratio, and the physical state (solid, liquid). The removal of 80% of ANF levels and recovery of 90% of proteins were achieved using solid sulfated polysaccharides. Large amounts of particle complexes were observed for sulfated polysaccharides as compared to carboxylated polysaccharides by confocal laser scanning microscopy. A 100-kDa polyethersulfone ultrafiltration membrane was effective in recovering proteins (80%) from ι-carrageenan/SWP complexes, and a small amount of loss of the polysaccharides was observed for each cycle.
A highly selective procedure to extract thiol-containing peptides (TCPs) from complicated soy glycinin hydrolysates (SGHs) was described. This procedure included the reduction of disulfide bonds by 1,4-dithiothreitol (DTT) and enrichment of TCPs through Thiopropyl-Sephrose 6B covalent chromatography. TCPs were confirmed using a strategy based on mass shift after differential alkylation of sulfhydryl groups with iodoacetamide and N-ethylmaleimide by matrix-assisted laser desorption ionization mass spectrometry (MALDI-TOF-MS). The antioxidant activities of TCPs were evaluated using chemical assays. DTT reduction increased the concentration of sulfhydryl groups from 1.8 μmol/g to 113.8 μmol/g. The efficiency of the extraction was improved by optimizing the loading of sample, extraction and desorption time and the content of desorption reagent. Both of the adsorption and desorption process were found to fit a pseudo-second order model. MALDI-TOF-MS showed that 36 of the 45 extracted peptides were TCPs. The EC50 of TCPs for DPPH, hydroxyl radical, and superoxide anion radical was 0.1, 1.49 and 0.084 mg/mL, respectively. The reducing power of TCPs (0.2 mg/mL) was of 0.375. These results suggest that the combination of DTT reduction and Thiopropyl-Sepharose 6B covalent chromatograph was a successful pathway to extract TCPs from SGHs and the TCPs could be used as potential antioxidants.
Effects of removal of non-network proteins by diffusion process on the storage modulus and loss modulus of soy protein isolate (SPI) gels and glycinin (11S) gels as a function of heating temperature and ionic strength were investigated. The composition of non-network proteins was determined by native-PAGE, non-reducing SDS-PAGE and reducing diagonal SDS–PAGE. Results showed that non-network proteins were composed of a majority of acid polypeptides (A), small amounts of AB subunits, soybean agglutinin, Bowman−Birk trypsin inhibitor, and lower amounts of α′, α and A3 polypeptides. The results further revealed that 11S gels had higher ratios of non-network proteins than SPI gels, due to the increased ratio of A polypeptides in 11S gel. In addition, the removal of non-network proteins from the gel was found to have no effect on the storage modulus, but on the other hand resulted in a decrease in the loss modulus, suggesting that the loss modulus of the gel network is closely related to non-network proteins. This study presents an approach to investigate the changes of storage modulus and loss modulus of globular gels in relation to the removal of non-network proteins, and provides valuable information on the composition and content of these proteins in gel network formed at various conditions.
Compared to other processing methods used in soybeans, roasting has received limited attention in soymilk processing despite its known potential role in improving soymilk quality. In this study, the effect of soybean roasting at 110 °C for 20 to 100 min and at 120 °C for 20 min on the distribution of soymilk volatile compounds, antinutritional factors and storage parameters: pH, total titratable acidity, emulsion stability (ES) and rheology were investigated and comparison was made with the control sample prepared from soybeans which were not roasted. Results have shown that roasting soybeans improved soymilk properties by significantly decreasing the levels of each of the following objectionable volatile compounds: hexanal, 1-hexanol and Furan, 2-pentyl respectively from 43.58% to a minimum level of 17.95, 11.67–2.28 and 4.09–0.82%. In addition, the levels of trypsin inhibitors were considerably decreased from 5.86 to 1.91 mg/gm thus reducing the overall heat treatment time of the soymilk from 19.4 to 9.9 min. ES increased significantly from 23.97 to 102.84 min while viscosity at the shear rates of 0.1 and 1.0/s decreased respectively from 3.00 and 0.73 to 0.27 and 0.03 Pa s. Furthermore, all the samples were found with the behavior flow indices values of less than unity (0.16–0.38 respectively) suggesting that they were pseudo plastics. From the results, it can therefore be concluded that careful selection and application of soybean roasting parameters in terms of temperature and time can be a useful strategy in improving the quality of soymilk.
We first observed that protein/polysaccharide interaction exhibited non-interacting behavior which makes Bowman-Birk chymotrypsin inhibitor (BBI) always free of complexation, being separated from another protein with similar isoelectric points, Kunitz trypsin inhibitor (KTI). Turbidity titrations showed that, the electrostatic attractions were much stronger between KTI/BBI (KBi) and carboxymethyl cellulose of higher substitution degree. Unchanged chymotrypsin inhibitory activity (CIA) indicated that BBI had negligible contribution to protein recovery and trypsin inhibitory activity (TIA). Tricine–SDS-PAGE revealed that at r = 20:1-2:1, unbound BBI was left in the supernatant when bound KTI transferred into precipitates, even if there was excess negative charges. Thus, purified KTI or BBI was achieved easily at the given conditions. The non-interacting behavior of BBI was further confirmed by ITC, where the binding enthalpy of BBI to CMC was negligible compared with the high binding affinity (Kb) of KTI. This work will be beneficial to protein purification based on protein–polysaccharide coacervation.
Wheat gliadin nanoparticles were prepared by antisolvent precipitation, and gum arabic (GA) was added to improve the stability of the nanoparticles. Effect of pH, gliadin/GA ratio and gliadin concentration on the gliadin-GA particles were studied by turbidimetric measurement. Spinodal lines for the GA-gliadin-solvent system were built under different pH. The influences of gliadin/GA ratio and their concentrations were represented in the diagram, thus the conditions for preparing stable composite were defined. The stability of GA-coated gliadin nanoparticles (Gliadin/GA = 1:3) was further studied compared with uncoated gliadin particles. GA-coated nanoparticles had a relatively good stability at pH 4.0–7.0 and remained a relatively low particle size with the elevated ionic strengths. They also had good thermal stability at 80 °C. Moreover, the interactions between gliadin particles and GA under different pH were further investigated. Hydrogen bonding was found to be the predominant force at pH 5.0, while hydrophobic force took charge of the formation of complex at pH 7.0. The findings are of great importance for extending the current knowledge about gliadin nanoparticles coupled with polysaccharides, thus providing valuable information for the development of gliadin-polysaccharide nanoparticles as potential nano-delivery systems.
Peanut seeds are rich in oil, which exists as oil bodies (OBs). By extraction, peanut crude OBs are obtained and can be used as a food ingredient. In a previous study, it was found that the crude OBs contained an unknown protease, which hydrolyzed the oleosins. This would disrupt the integrity of OBs, and therefore, affect their physical and oxidative stability. In this study, the protein composition of crude OBs and some properties of the unknown protease were examined. The results showed that the protease was a two-chain (32 and 9kDa) aspartic protease, which showed high affinity for OBs. The optimal pH and temperature for oleosin hydrolysis by the protease were pH 4.0 and 60°C. Interestingly, the aspartic protease not only hydrolyzed OB intrinsic proteins (oleosin, caleosin, and steroleosin), but also extrinsic proteins (especially Ara h 1 allergen and 26-30kDa arachin).
Emulsifying behaviors and interfacial properties were two key properties related to microencapsulation processes. This study mainly focused on the above characters of soy proteins/gum arabic (SP/GA) complexes and gelatin/GA complexes. The complexes were prepared at different forming stages (i.e., at pHφ1, pHopt and pHpd) with 1:1 protein/GA ratio. The results revealed the emulsifying activity (EA) for SP/GA systems was higher than that for gelatin/GA systems, indicating its better combining capacity with oil droplets. While SP/GA systems showed lower interfacial tensions (21–25 mN/m) than gelatin/GA systems. Through analysis of interfacial tensions and EA data, negative correlations between them were found: the higher the interfacial tensions of complexes with oil droplet, the lower the EA was. Compositional analysis indicated different yield and polymer recovery were obtained at different systems due to ζ potential differences. These findings suggested SP/GA complexes could be used as effective microencapsulated materials in terms of oil combining.
It is generally acknowledged that despite the superior nutritional value of soymilk, the presence of off-flavours catalysed by lipoxygenase (LOX) activities has to some extent negatively influenced the overall acceptability of soymilk in consumers. In this current study, the effect of roasting soybeans at different temperatures (110–130 °C) and times (20–120 min) on physical, chemical properties and LOX activity of soymilk was investigated in order to establish optimum soybean roasting conditions for developing nutritious and LOX free soymilk. Results have revealed that subjecting soybeans to increased roasting temperature and time negatively affected physical chemical properties of soymilk by decreasing the yield, solid content, colour and nutritive value. On the other hand, roasting soybeans at a relatively low temperature of 110 °C for 80 min significantly inactivated LOX but retained adequate protein content in soymilk. The findings of this study have demonstrated that careful selection of roasting conditions can be a useful strategy for inactivating LOX activity in development of a more acceptable soymilk product.
P34 probable thiol protease (P34) and Gly m Bd 30K (30K) show high relationship with the protease of 24 kDa oleosin of soybean oil bodies. In this study, nine days germinated soybean was used to separate bio-processed P34 (P32) from bio-processed 30K (28K). Interestingly, P32 existed as dimer, whereas 28K as monomer; P32-rich sample had proteolytic activity and high cleavage site specificity (Lys-Thr of 24 kDa oleosin), whereas 28K-rich sample showed low proteolytic activity; P32-rich sample contained one thiol protease. After mixing with purified oil bodies, all P32 dimers were dissociated and bound to 24 kDa oleosins to form P32-24 kDa oleosin complexes. By incubation, 24 kDa oleosin was preferentially hydrolyzed, and two hydrolyzed products (HPs; 17 and 7 kDa) were confirmed. After most of 24 kDa oleosin was hydrolyzed, some P32 existed as dimer, and the other as P32-17 kDa HP. It was suggested that P32 was the protease.
The properties of oil-in-water emulsions containing soy protein isolate (SPI) with or without gum arabic (GA) have been studied under different pH and different preparation routes. Three routes were used for the preparation of emulsions (10% (w/v) oil and 0.5% (w/v) SPI with or without 0.5% (w/v) GA). In emulsions I, SPI solution with/without GA were first pH adjusted followed by homogenization with oil; in emulsions II, the oil was directly emulsified with SPI–GA coacervates at pH 4.0 to form the ‘mixed emulsions’, then pH was adjusted; in emulsions III, GA was added to a SPI-stabilized emulsion followed by pH adjustment. The results showed that pH adjustment before or after homogenization with oil greatly influenced the droplet size and emulsifying stability whether for the SPI emulsions or SPI/GA emulsions. The emulsions III showed much slower flocculation rate and higher emulsifying stability at pH 4.0–7.0. The microstructures of SPI–GA emulsion III were more uniform, with the comparably smallest d43 values in the presence of SDS. It could be indicated that the preparation routes of emulsions greatly influenced the stability of SPI–control emulsion or SPI–GA emulsion.
The diffusion of poly(ethylene glycol) (PEG) in transglutaminase (TGase) cross-linked soya protein gels as affected by enzyme dosage and protein composition was investigated. PEG with different molecular weights (Mw) varying from 6000 to 400 000 g mol−1 was selected. Results of this study revealed that the time-related release of PEG from gel slices was found to be controlled by Fickian diffusion process. For PEG with Mw 6000 g mol−1, the diffusion coefficients (D) were independent on the enzyme dosage and protein composition. However, for PEG with Mw ≥ 20 000 g mol−1, there were strong dependences of diffusion on the solute size and on the gel network microstructure revealed by scanning electron microscopy (SEM), where more density gel network with smaller pores resulted in lower D of PEG. Lower D values of PEG with Mw ≥ 20 000 g mol−1 were obtained in gels induced by 2 U g−1 TGase, and the D values increased with increasing 11S ratio in gels. Moreover, data also showed that the diffusion of PEG with larger size was more sensitive to the variation of gel network.
Due to the complications of the soymilk system, the heat-induced Bowman-Birk inhibitor (BBI) inactivation mechanism is not well known. In this study, two BBI samples with low and high purities were prepared from soymilk. It was confirmed that three groups (A, C, and D) of BBI, which are contained in soybean seeds, were transferred into soymilk during processing. On heating, it was found that 1) the two subdomains of BBI were not equally heat stable, 2) the conformation of BBI gradually changed, 3) some amino acid residues (namely, cystine, serine and lysine) in BBI were degraded, 4) BBI did not tend to form intermolecular cross-links with another BBI, but did slightly with non-BBI proteins. Based on some previous studies, the conformational change of BBI was attributed to β-elimination reactions on the amino acid residues of BBI and the subsequent intramolecular reactions induced by the products yielded by the β-elimination reactions.
Soybean seeds contain three groups (A, C, and D) of Bowman-Birk inhibitors (BBI). In this study, highly purified BBI-A (approximately 96%) was obtained from soybean whey at the 0.1 g level by the complex coacervation method. BBI-A has seven disulfide bonds (SS) and no sulfhydryl group and exhibits trypsin inhibitor activity (TIA) and chymotrypsin inhibitor activity (CIA). The X-ray structure has shown that BBI-A has five exposed SS and two buried SS. Due to steric hindrance, it was reasonable to consider that dithiothreitol first attacks the five exposed SS and then the two buried SS, which was supported by the results that SS reduction with dithiothreitol could be divided into quick and slow stages, and the critical point was close to 5/7. The effects of SS reduction on TIA and CIA could be divided into three stages: when one exposed SS was reduced, both TIA and CIA decreased to approximately 60%; with increasing reduction of exposed SS, CIA gradually decreased to 8%, and TIA gradually decreased to 26%; with further reduction of buried SS, CIA gradually decreased to 2%, and TIA slightly decreased to 24%. Far-UV CD spectra showed that the secondary structure of BBI-A was slightly changed, whereas near-UV CD spectra showed that the conformation of BBI-A was substantially changed after the five exposed SS were reduced, and further reduction of buried SS affected the conformation to some extent. The results from Tricine-SDS-PAGE and C8 column showed the same trend as near-UV CD spectra. BBI-A has a structural peculiarity in that two hydrophobic patches are exposed to the exterior (in contrast to typical soluble proteins), which was attributed to the seven SS by some researchers. The results above supported the hypothesis that hydrophobic collapse of the exposed hydrophobic patches into a regular hydrophobic core occurred after the reduction of SS in BBI-A.
Two successive and selective coacervations induced by chitosan (Ch) and carrageenan (CG) were applied to remove antinutritional protease inhibitors and purify Bowman–Birk protease inhibitor (BBI) from soybean whey. At the first coacervation induced by Ch (66.7, 200, and 510 kDa), only Kunitz trypsin inhibitor (KTI) and BBI complexed with Ch were extracted, while β-amylase and soybean agglutinin remained in supernatant. The binding constants for the interaction increased on the order Ch-66.7 < Ch-200 < Ch-510. At the second selective complexation, we observed a competitive binding behavior between KTI/BBI and CG. At a mixing weight ratio of 3:1 (pH 3.0 for ι-CG, and pH 3.11 for λ-CG), the preferential binding of KTI to CG led to the single enrichment of BBI in the supernatant. Our results indicated that the purified BBI was a good source for further study of its anti-carcinogenic properties, due to its high bioactivity (669.5 U/mg chymotrypsin-inhibitory activity and 2260 U/mg trypsin-inhibitory activity).
In this study, the effect of roasting temperatures (110, 120 and 130o C) and times (20, 40, 60, 80, 100 and 120 min) respectively on soybean protein quality test parameters was investigated. Results have revealed that urease activity decreased gradually with increasing time at 110C, suggesting that the test could be a suitable indicator for both under processing and over processing as opposed to 120 and 130C where there was a sudden drop in urease activity. Results further showed that at 110C, protein solubility in potassium hydroxide remained high with increasing time while at 120C, protein solubility decreased inconsistently. On the other hand, protein solubility at 130C decreased steadily suggesting that the test could be a suitable indicator for both under processing and over processing. It was further observed that at all roasting temperatures, protein dispersibility index decreased gradually with the highest and lowest decreases observed at 130C and 110C respectively. Results further showed that at 130C, protein digestibility and protein dispersibility index tests could yield results that were comparable with urease activity and protein solubility tests unlike at 110 and 120C. The findings have demonstrated that roasting temperatures and times significantly affected the test parameters used in determining the adequacy of soybean processing. These findings justify the need to carefully consider roasting temperatures for potential applications of processed soybeans in animal feeds processing as well as product development for human consumption.
The effects of heating temperature, ionic strength and 11S ratio on the rheological properties of heat-induced soy protein gels, particularly in relation to network proteins content and aggregates size were investigated. Results revealed that non-network proteins consisted of undenatured glycinin (11S) AB subunits, high levels of A and A3 polypeptides but low levels of β-conglycnin (7S) α and α′ subunits. Results further showed that a positive correlation between storage modulus (G′) and network proteins ratio of gels that were formed at different temperatures. Additionally, the G′ values of gels initially increased with increasing NaCl concentration from 0 to 0.25 M, but decreased with further increasing ionic strength from 0.25 to 0.50 M. This trend was consistent with the variation of heat-induced protein aggregates size in soy protein solution at the same ionic strength. Higher 11S/7S ratio resulted in higher storage modulus values of soy protein gels due to the formation of larger and compacter aggregates mainly by B polypeptides via hydrophobic bonds.
The effect of polymer charge density on protein selectivity in the presence of carboxylated polysaccharides (CPS) and sulfated polysaccharides (SPS) was investigated for Kunitz trypsin inhibitor/Bowman-Birk protease inhibitor (KTI/BBI, KBM). To determine the conditions for coacervation or precipitation as a function of polymer charge densities, turbidimetric titrations and Tricine–SDS-PAGE were used. Polymer charge density as well as chain flexibility greatly influenced the strength of interactions and protein recovery. While charge compensation must occur for CPS–KBM complexes, SPS–KBM systems did not require conservation of charge neutrality. Despite their similar isoelectric points, KTI bound preferentially to CPS and SPS due to its higher affinity compared to BBI. Complexation of KBM with the polysaccharide with the lowest charge density, arabic gum expectedly cannot realize the purification of BBI under conditions where binding to more highly charged polysaccharides occurs. This work will be beneficial to selective purification of target proteins through control of protein–polysaccharide complexation.
The release behavior of poly(ethylene glycol) (PEG) with different molecular weights (Mw): 6000, 20 000, 100 000 and 400 000 g mol⁻¹ from heat-induced soy protein gels as a function of the 7S/11S protein ratio was investigated. Results showed that time-related release of PEG from gel network fit the Fick's second law. PEG diffusion coefficients depended on the PEG size and gel network structure, which increased with increasing 11S ratio in gel or decreasing probe molecular weight. Moreover, the diffusion of PEG with higher molecular weight was more sensitive to the variations of gel internal network. The changes of PEG diffusion coefficients in soy protein gel were consistent with the variations of gel structure characterized by scanning electron microscopy (SEM). Gels formed at lower 11S ratio were composed of homogeneous spherical aggregates structure, while gels formed at higher 11S ratio showed higher extent of macro-phase separation and coarser network with larger pores.
The interaction between glycinin and anionic polysaccharides has gained considerable attention recently because of its scientific impact on the stability of acid soymilk systems. In this study, the formation of glycinin/gum arabic complexes driven by electrostatic interactions was investigated. Turbidity titrations at different glycinin/gum arabic ratios were conducted and critical pH values (pHφ1) where insoluble complexes began forming were determined firstly. The corresponding pHφ1 values at glycinin/gum arabic ratios of 1:4, 1:2, 1:1, 2:1, 4:1 and 8:1 were 2.85, 3.25, 3.70, 4.40, 4.85 and 5.35, respectively. Afterwards, electromobilities for glycinin and gum arabic at the pH values between 4.1 and 2.6 were measured, and charge densities (ZN) for glycinin and gum arabic were calculated based on the soft particle analysis theory. Further analysis indicated that the product of glycinin/gum arabic ratio (ρ) and ZN ratio of glycinin/gum arabic was approximate 1 at any pHφ1 values. It was revealed that charge neutralization was achieved when glycinin/gum arabic insoluble complexes began forming. NaCl displayed multiple effects on glycinin/gum arabic complex formation according to turbidity and compositional analysis. The present study could provide basic guidance in acid soymilk designing.
Complex behavior of Bowman–Birk protease inhibitor (BBI) with ι-carrageenan (LC) as a function of pH, protein to polysaccharides ratio and salt concentration was studied by turbidimetric titration, dynamic light scattering (DLS) and isothermal titration calorimetry (ITC). At fixed BBI/LC weight ratio of 5:1, turbidity and DLS results showed that pHc and pHφ1 shift to the lower pH values with the increase in ionic strength (I), whereas the former occurred at the pH lower than isoelectric point (pI = 4.2) of BBI at I ≥ 100 mM NaCl. ITC results showed that BBI binding to LC involves a two-step process with an increasing exothermic enthalpy at the first binding step. The further insight of BBI-LC complexation was studied as a function of BBI concentration, ionic strength and temperatures using ITC. The critical molar ratio (rcritical) between two binding steps was independent of protein concentration, ionic strength and temperature, although the heat flow obviously decreased with the increasing I (0–200 mM) and slightly increased with the elevated temperature (25–45 °C). The negative heat capacity (ΔCp) and the gain in nonionic contribution (ΔGno) indicated the involvement of nonelectrostatic interactions (e.g., hydrophobic effect) for the first binding step.
Soybean oil bodies (OBs), naturally pre-emulsified soybean oil, have great potentials to be used in foods and cosmetics. In this study, OBs were recovered from soybean aqueous extract at pH 6.8, 8.0, 9.5, and 11.0, and recovered OBs contained decreased extrinsic protein amount and composition with increasing recovery pH. In unheated condition, particle size and viscosity decreased, whereas isoelectric point (pI) and oxidative stability increased in the order of pH 6.8-, 8.0-, 9.5-, and 11.0-OB. By heating, it was observed that 1) coalescence of OBs occurred in pH 6.8-OB emulsion, but not occurred in pH 11.0-OB emulsion; 2) pIs of all OB emulsions increased; 3) pH 9.5-OB emulsion showed the highest viscosity, followed by pH 8.0- and 6.8-OB, and pH 11.0-OB still showed the lowest viscosity; 4) gels were formed from OB emulsions with solid content of 40% except pH 11.0-OB; 5) oxidative stability was greatly improved for all OB emulsions. This study is meaningful for supplying fundamental information for selecting proper conditions for aqueous extraction of soybean OBs.
Plant seeds are used to extract oil bodies for diverse applications, but oil bodies extracted at different pH values exhibit different properties. Jicama, sunflower, peanut, castor bean, rapeseed, and sesame were selected to examine the effects of pH (6.5–11.0) on the protein components of oil bodies and the oleosin hydrolysis in pH 6.5-extracted oil bodies. In addition to oleosins, many extrinsic proteins (globulins, 2S albumin, and enzymes) were present in pH 6.5-extracted oil bodies. Globulins were mostly removed at pH 8.0, whereas 2S albumins were removed at pH 11.0. At pH 11.0, highly purified oil bodies were obtained from jicama, sunflower, peanut, and sesame, whereas lipase remained in the castor bean oil bodies and many enzymes in the rapeseed oil bodies. Endogenous protease-induced hydrolysis of oleosins occurred in all selected plant seeds. Oleosins with larger sizes were hydrolysed more quickly than oleosins with smaller sizes in each plant seed.
Proteins in soybean whey were separated into two groups by graded salt precipitation, low and high isoelectric point protein fractions (LIP and HIP), corresponding to the mixture of Kunitz trypsin inhibitor (KTI) and Bowman-Birk protease inhibitor (BBI) as well as the mixture of soybean agglutinin (SBA) and β-amylase, respectively. The complex behavior of LIP and HIP with ι-carrageenan (CG) as a function of pH (7.0-2.0) and protein-polysaccharide mass mixing ratio (1:1 to 20:1, w/w) was studied by turbidimetric titration and SDS-PAGE. During pH titration, pHφ1 (the initial pH for the formation of insoluble complexes) and pHmax (the pH for maximum optical density) showed mixing ratio dependence so that the choice of mixing ratio determined the order of coacervation. SDS-PAGE results showed that at protein/polysaccharides mass ratio (Rpr/ps) of ≥15:1, just only KTI (or SBA) complex with ι-carrageenan and shift to precipitates after centrifugation, BBI (or β-amylase) was kept in supernatant. This selectivity was also independent of the total biopolymer concentration (Cp) and titration path. After removal of carrageenan, high-purity KTI, BBI and SBA (over 90% by SEC-HPLC) were obtained. Isothermal titration calorimetry (ITC) showed that, the smaller thermodynamic stoichiometry (n) with higher thermodynamic constants (K) was observed when KTI (or SBA) complexing with CG compared to BBI (or β-amylase). The higher surface charge density of KTI (+0.22 × 10-2 C/m2) and SBA (+0.14 × 10-2 C/m2) may contribute partly to their higher CG thermodynamic affinity.
In unheated soymilk, extrinsic proteins are bound to intact oil bodies coated by one monolayer of phospholipids and oil body intrinsic oleosins. In this study, effects of heating (70-100 °C; 0-30 min) on particle size and bound proteins of oil bodies were examined in suspension (oil bodies from unheated soymilk into deionized water) and soymilk. Mass ratio of extrinsic proteins/oleosins of oil bodies in unheated suspension and soymilk was respectively 1.1 and 2.5. By heating, extrinsic proteins released from oil bodies with different rates, and Z-average size of oil bodies increased in the beginning; afterwards, residual extrinsic proteins (extrinsic proteins/oleosins: 0.31 in suspension and 0.74 in soymilk; 100 °C, 4 min) and Z-average size were affected little. The amount of residual proteins at 80-100 °C was negatively correlated with Z-average size of oil bodies in suspension (R2 = 0.996) and soymilk (R2 = 0.890).
The effects of heat treatment on the emulsifying properties of pea proteins were investigated. Thermal treatment of pea proteins at 95 °C for 30 min increased the extent of protein aggregation, and the hydrodynamic diameter increased with the increasing of heated protein concentration (ch). Electrophoresis showed that acidic and basic (AB) subunits as well as convicilin in unheated pea proteins were involved in the formation of polymers linked by disulfide bonds (SS) after heat treatment (95 °C, 30 min). At different protein concentrations in the continuous phase (c: 0.1%-0.5%, w/v) with constant oil fraction of 0.1, emulsions formed by heated pea proteins (95 °C, 30 min) showed higher protein adsorption percentage and creaming stability than those formed by unheated proteins. Proteins adsorbed at the oil-water interface contained higher percentages of vicilin and basic subunit of legumin (leg B) in emulsions stabilized by heated pea proteins than in those stabilized by unheated proteins. Moreover, increasing c was conducive to the formation of emulsions with greater stability against creaming. In addition, emulsion viscosity increased with the increasing of ch. These results indicated that the heated pea proteins, as compared to the unheated pea proteins, exhibited a greater potential to act as a kind of excellent emulsifiers.
Development of viable alternatives to antibiotics to control necrotic enteritis (NE) caused by Clostridium perfringens is becoming urgent for chicken production due to pessures on poultry producers to limit or stop the use of antibiotics in feed. We have previously identified citral as a potential alternative to antibiotics. Citral has strong antimicrobial activity and can be encasupsulated in a powder form for protection from loss during feed processing, storage, and intestinal delivery. In the present study, encapsulated citral was evaluated both in vitro and in vivo for its antimicrobial activity against C. perfringens. Encapsulation did not adversely affect the antimicrobial activity of citral. In addition, encapsulated citral was superior to the unencapsulated form in retaining its antimicrobial activity after treatment with simulated gastrointestinal fluids and in the presence of chicken intestinal digesta. In addition, the higher antimicrobial activity of encapsulated citral was confirmed in digesta samples from broilers that had been gavaged with encapsulated or unencapsulated citral. In broilers infected with C. perfringens, the diets supplemented with encapsualted citral at both 250 and 650 μg/g significantly reduced intestinal NE lesions, which was comparable to the effect of bacitracin- and salinomycin-containing diets. However, supplementation with the encapsulated citral appeared to have no significant impact on the intestinal burden of Lactobacillus. These data indicate that citral can be used to control NE in chickens after proper protection by encapsulation.
This study aimed to develop an optimal continuous procedure of immobilized hydroperoxide lyase (HPL)-catalyzed synthesis of hexanal. A central composite design was used to study the combined effect of substrate concentration and the residence time of the reactant on hexanal concentration. The optimum conditions for hexanal synthesis included a 13-HPOD concentration of 43.54 mM and a residence time of 60.99 min. The maximum hexanal concentration was 3560 ± 130 mg/L when 16 U of immobilized HPLwas used. Furthermore, the stability of immobilized HPL was significantly improved in the packed-bed reactor, as evidenced by the slowed enzyme inactivation and prolonged operation time. The immobilized HPL remained activity until 40 mL substrate solution flowed past the packed-bed reactor. The catalyst productivity of hexanal in the packed-bed reactor was 5.35 ± 0.34 mg/U, much higher than that in the batch stirred reactor. This study was greatly meaningful for providing a green method to the large-scale production of hexanal.
A simple and efficient method for preparing Tricine-SDS-PAGE protein sample of extracted oil bodies (OBs) was supplied: OB suspension was vortexed with SDS buffer (pH 6.8) for 2min at room temperature with SDS/protein of 1.52/1(w/w), which could be analyzed by Tricine-SDS-PAGE after simple treatments (dilution and 2-mercaptoethanol). At SDS/protein of 1.52/1, about 95% of proteins in soybean OB suspension were solubilized, whereas residual 5% of proteins were weakly bound to SDS-destroyed OBs; proteins in destroyed OBs might be further solubilized by SDS in the gel and cathode buffer of Tricine-SDS-PAGE, causing about 99% of proteins in soybean OB suspension recover on Tricine-SDS-PAGE gel, which was better than acetone (89%) and diethyl ether (96%) harvested protein samples. Higher or lower SDS/protein was unbeneficial for protein solubilization from OBs. Additionally, the above method was also better than organic solvent method for peanut, sesame, and rapeseed OB suspensions. Copyright © 2015 Elsevier Ltd. All rights reserved.
Potato proteins (PP) in the lab-made potato juice consists of patatins (38.1%), protease inhibitors (56.7%) and other high molecular weight proteins (5.1%) as characterised by Tricine-SDS-PAGE and MALDI-TOF/TOF-MS. To recover PP from potato juice, the complex behaviours of PP with several natural polyelectrolytes as a function of pH and protein to polyelectrolyte ratio (RPP/Polysaccharide) were studied by turbidimetric titration. Results indicated that the turbidity curves of PP-polysaccharides displayed a shift to higher pH for chitosan and lower pH for anionic polysaccharides with the decreasing of RPP/Polysaccharide. Chitosan could be used to selectively recover patatin with the purity of 88.6%, and the highest protein yield (51.9%) was achieved at pH 6.0 and RPP/chitosan of 1:2. Carrageenan, one typical anionic polysaccharide, could recover all the PP in potato juice at pH 3.5 and RPP/carrageenan of 2.5:1 with no special protein selectivity.
The processing parameters for making a Maillard reaction product (SPPMP) from soy protein isolate (SPI) and soy soluble polysaccharide (SSPS) were studied against the yield of the product and its emulsification capacity in an oil-in-water emulsion. The optimized SPPMP was produced by dry-heating the SPI-SSPS mixture (SPP) at a ratio of 3:5, temperature of 60 °C and 75% relative humidity for 3 days. The formation of SPI-SSPS conjugates was confirmed by gel electrophoresis, FTIR spectroscopy and high performance size exclusion chromatography. The citral (10 wt%) oil-in-water emulsions stabilized by SPPMP exhibited superior physical stability than those stabilized by SPI or SPP during prolonged storage, after thermal treatment or under simulated gastrointestinal conditions. At pH 7.0, all the emulsions studied exhibited monomodal particle size distribution initially, however, only those stabilized by SPPMP remained monomodal distribution for up to 70 days during storage at 25 °C. The SPPMP-stabilized emulsion maintained its physical stability to the thermal treatment at 95 °C for 30 min or under simulated gastric conditions for 2 h; while the emulsions stabilized by SPI or SPP exhibited various degrees of instability. The release rate of citral from the emulsion droplets was found inversely related to the stability of emulsion. The emulsion droplets retained approximately 70% of citral after 2 h incubation in simulated gastric fluid, whereas, complete release of citral from the droplets occurred in 4 h in simulated intestinal fluid. These results indicate that SPPMP-stabilized emulsions have a good potential as a carrier system for intestinal delivery of hydrophobic compounds such as citral.
Application of soya proteins (SP) in flaxseed oil microencapsulation based on complex coacervation was investigated. The effects of SP/gum arabic (GA) mixing ratio (1:2, 1:1 and 2:1) and pH (2.80, 3.15 and 3.75) on coacervate preparation were studied firstly. The highest coacervate yields (CY) were achieved at SP/GA = 1:1, pH 3.15, and SP/GA = 2:1, pH 3.75, which were 81.2 ± 2.0%, 88.1 ± 0.6%, respectively. Thereafter, the microencapsulation of flaxseed oil was detected in accordance with the results of the coacervation of SP/GA, that is the optimal condition for microencapsulation was corresponded to the condition where CY was the highest. Under the conditions of SP/GA = 1:1, pH 3.15, and SP/GA = 2:1, pH 3.75, the microencapsulation efficiency and total yield reached 81.5 ± 0.1% and 81.7 ± 0.4%, and 77.4 ± 3.7% and 86.7 ± 2.4%, respectively. Microscopic morphology revealed that the formation of a biopolymer shell around the oil droplets was achieved at specific conditions.
Complexing between soy proteins (SP) and gum arabic (GA) was achieved by mutual titration of soy protein and gum arabic and was characterized using isothermal titration calorimetry (ITC), turbidity, sedimentation and ternary phase boundaries. In the first section, SP were titrated into GA (SP-to-GA titration) under salt-free condition (no added NaCl) at pH 3.0 and pH 5.6, respectively. ITC experiments displayed exothermic processes at both pH status, but the enthalpy changes (ΔH) at pH 3.0 was −0.70 ± 0.02 cal/g as compared to −0.10 ± 0.01 cal/g at pH 5.6. For SP-to-GA titration at pH 3.0, a sudden turbidity increase was observed at the critical SP/GA mass ratio (rφ) of 0.42, which was approximately equal to the charge density ratio of GA and SP (0.36), indicating the charge compensation was achieved at phase separation point. In the second part, GA was titrated into SP (GA-to-SP titration) under salt-free condition at pH 3.0. An immediate turbidity increase was observed when GA was added into SP, while the sedimentation ratio measurement showed that the complex was unstable only in the GA/SP mass ratio range of 0.3–0.6. The ITC result showed a much higher ΔH than that for SP-to-GA titration. Effect of NaCl concentration on the complexing behavior by SP-to-GA titration at pH 3.0 was studied in the last. ΔHs and binding isotherms changed monotonically with the increase of salt concentration from 0 to 250 mM. However, turbidity measurement and phase boundaries revealed that the maximum phase separation was obtained at salt concentration of 100 mM.
Heat-induced soy protein gels were prepared by heating protein solutions at 12%, 15% or 18% for 0.5, 1.0 or 2.0 h. The release of non-network proteins from gel slices was conducted in 10 mM pH 7.0 sodium phosphate buffer. SDS-PAGE and diagonal electrophoresis demonstrated that the released proteins consisted of undenatured AB subunits and denatured proteins including monomers of A polypeptides, disulfide bond linked dimers, trimers and polymers of A polypeptides, and an unidentified 15 kDa protein. SEC-HPLC analysis of non-network proteins revealed three major protein peaks, with molecular weights of approximately 253.9 kDa, 44.8 kDa and 9.7 kDa. The experimental data showed that the time-dependent release of the three fractions from soy protein gels fit Fick's second law. An increasing protein concentration or heating time resulted in a decrease in diffusion coefficients of non-network proteins. A power law expression was used to describe the relationship between non-network proteins diffusion coefficient and molecular weight, of which the exponent (α) shifted to higher value with an increase in protein concentration or heating time, indicating that a more compact gel structure was formed.
Many thiol-containing molecules show heavy metal complexation ability and are used as antidotes. In this study, the potential function associated with thiol-containing peptides (TCPs) from soy protein hydrolysates as natural detoxicants for heavy metals is reported. TCPs enriched by Thiopropyl-Sepharose 6B covalent chromatography had different molecular weight distributions as well as different numbers of proton dissociable groups, depending on the proteases and degree of hydrolysis. The major contribution of sulfhydryl groups was confirmed by the largest pH decrease between 8.0 and 8.5 of the pH titration curves. The complexation of TCPs with heavy metals was evaluated by stability constants (βn) of TCP-metal complexes whose stoichiometry was found to be 1:1 (ML) and 1:2 (ML2). TCPs from degree of hydrolysis of 25% hydrolysates gave high affinities towards Hg2+, Cd2+, and Pb2+ (giving similar or even bigger lgβ values than that of glutathione). A significantly positive correlation was found between the logarithm of stability constants for ML2 (lgβ2) and the sulfhydryl group content. Molecular weight distribution of TCPs affected the complexation with Pb2+ notably more than Hg2+ and Cd2+. These results suggest that soy TCPs have the potential to be used in the formulation of functional foods to counteract heavy metal accumulation in humans.
Citral is an important essential oil with antibacterial activities, but its use as an antibiotic alternative is limited due to its physical and chemical instability during processing and in biological systems such as the gastrointestinal tract of animals. This study aimed to investigate the capacity of a soy protein-polysaccharide Maillard reaction product (SPPMP) to stabilize citral in an oil-in-water emulsion system. The retention rates of citral in the emulsions during long time storage, upon heating and under simulated gastrointestinal conditions were determined. The results showed that SPPMP-stabilized emulsions demonstrated outstanding ability to stabilize citral under all challenge conditions as compared to emulsions stabilized by soy protein only, or by physical mixtures of soy protein and polysaccharide. Therefore, SPPMP-stabilized emulsions could potentially be used as protectors and carriers for targeted delivery of citral or other hydrophobic compounds to animal/human intestines.
It was well known that the disulfide-mediated interactions are important when soy protein are heated, in which soy proteins are dissociated and rearranged to some new forms. In this study, the disulfide bond (SS) linked polymer, which was composed of the acidic (A) polypeptides of glycinin, basic (B) polypeptides of glycinin, and a small amount of α' and α of β-conglycinin, was formed as major product, accompanied by formation of SS linked dimer of B and monomer of A as minor products. The role of sulfhydryl (SH) of different subunit/polypeptides for forming intermolecular SS was investigated. The SH of B in glycinin (Cys298 of G1, Cys289 of G2, Cys440 of G4) was transformed into SS in polymer identified by mass spectrometry analysis. The SH content of polymer was lower than that of glycinin and β-conglycinin subunits when heated. The SH content of B in polymer was lower than that in glycinin subunit, and both of them were decreased by heating. The SH content of A in polymer was increased and higher than that of B in polymer and A in glycinin subunit when heated. These results indicated that the SH of B in glycinin subunit was subjected to not only SH oxidation but also SH-SS exchange (with SS of A) for forming intermolecular SS of polymer. The SH oxidation and SH-SS exchange were proven by the change of SH content for the first time. The SH of B was suggested to be reactive for forming intermolecular SS of polymer.
During thermal treatment of soymilk, a rapid incorporation of Kunitz trypsin inhibitor (KTI) into protein aggregates by covalent (disulfide bond, SS) and/or noncovalent interactions with other proteins is responsible for its fast inactivation of trypsin inhibitor activity (TIA). In contrast, the slow cleavage of a single Bowman-Birk inhibitor (BBI) peptide bond is responsible for its slow inactivation of TIA and chymotrypsin inhibitor activity (CIA). In this study, the effects of Ohmic heating (220 V, 50 Hz) on soymilk TIA and CIA inactivation were examined and compared to induction cooker and electric stove heating with similar thermal histories. It was found that: (1) TIA and CIA inactivation was slower from 0 to 3 min, and faster after 3 min as compared to induction cooker and electric stove. (2) The thiol (SH) loss rate was slower from 0 to 3 min, and similar to induction cooker and electric stove after 3 min. (3) Ohmic heating slightly increased protein aggregate formation. (4) In addition to the cleavage of one BBI peptide bond, an additional reaction might occur to enhance BBI inactivation. (5) Ohmic heating was more energy-efficient for TIA and CIA inactivation. (6) TIA and CIA inactivation was accelerated with increasing electric voltage (110, 165, and 220 V) of Ohmic heating. It is likely that the enhanced inactivation of TIA by Ohmic heating is due to its combined electrochemical and thermal effects. © 2015 Institute of Food Technologists®
Coagulants and raw materials effects on the sensory characteristics of acid and salt-induced soft tofu-type gels were investigated and compared. Gels made from soybean were more whitish than the ones made from SPI with L∗ values between 79.743 and 80.847. Similarly, the raw materials affected more the volatile compounds composition than the coagulants. It was also established that CaSO4 produced softer gels with hardness values varying between 104 and 145 g compare to 173 and 302 g for acid-induced gels. Salt-induced soybean gels demonstrated the highest score for beany taste (7.67), bitter taste (7.39) and mouthfeel (7.25). Correlation results depicted that the volatile compounds did not have a positive nor negative significant impact on mouthfeel of soft tofu gels; however, they were significantly correlated to beany and bitter taste. Furthermore, all free amino acids were positively correlated to beany, bitter taste and mouthfeel except cysteine. Glutamic acid, alanine, tyrosine, methionine and isoleucine free amino acids showed a significant influence on beany taste; while, glutamic acid, tyrosine, methionine and isoleucine showed a significant impact on bitter taste.
Poly(γ-glutamic acid) (γ-PGA) is a natural, biodegradable polyamide that is mostly produced by glutamate-dependent micro-organisms such as Bacillus licheniformis, Bacillus amyloliquefaciens and Bacillus subtilis. In this study, a γ-PGA-producing glutamate-independent strain, SK19.001, was screened and characterized as Bacillus methylotrophicus. The effects of various carbon and nitrogen sources on the γ-PGA productivity of this strain were investigated. Many carbon sources, such as sodium glutamate, glucose, glycerol, soluble starch, α-lactose, maltose, sucrose, citric acid and fructose, proved suitable for SK19.001 to synthesize γ-PGA with high molecular weight. Compared with the previously reported production from glutamate-independent strains, a high level of production of γ-PGA (33.84-35.34 g/L) was obtained when SK19.001 was grown in a medium containing 30 g/L glycerol, 15 g/L sodium citrate and 50 g/L peptone. The γ-PGA product showed an ultra-high molecular weight of over 10,000 kDa. The genome of Bacillus methylotrophicus SK19.001 was sequenced and a comparative investigation revealed that the PgsB, PgsC, and PgsA protein sequences were almost identical between the glutamate-independent strains and less identical between the glutamate-independent and glutamate-dependent strains. This study is the first to demonstrate high levels of γ-PGA production using B. methylotrophicus. The results suggest a promising new approach for γ-PGA production.
We investigated the effects of rutin additive on the physical (droplet size, zeta potential, and interfacial adsorption) and oxidative stability of emulsions stabilized by native soybean protein isolate (SPI) and heat-denatured SPI. The fluorescence spectra and interfacial tension of SPI-rutin mixture were also measured. The formation of SPI-rutin complex mainly driven by hydrophobic interactions was verified. The addition of rutin reduced interfacial tension of native SPI at the oil-water interface, while that of HSP was less influenced. In the case of native SPI emulsion, rutin particles could competitively adsorb into the oil-droplet interface with high interfacial accumulation. The formation of native SPI-rutin mixed interfacial layers resulted in the reductions in oil droplet size of emulsions and evident improvement of physical and oxidative stability. However, the stability of heated SPI emulsions showed no remarkable improvement due to lower interfacial accumulation and physical location of rutin. Rutin particles were believed to mainly adsorb outside protein layer. These findings showed that rutin can be used as a native stabilizer for protein emulsions to enhance its physical and oxidative stability.
Phytate is an important anti-nutritional factor in food products. In this study, phytase-assisted processing method was used to produce low-phytate soybean protein isolate (SPI) samples, and their physicochemical and functional properties were examined. Hydrolysis condition at low temperature (room temperature) and pH 5.0 was better than that recommended by manufacturer (pH 5.0, 55 °C) at keeping the properties of SPI, so the former condition was selected to prepare SPI samples with phytate contents of 19.86-0.11 mg/g by prolonging hydrolysis time (0 (traditional method), 5, 10, 20, 40, and 60 min). Ash content (R2 = 0.940), solubility (R2 = 0.983), ζ-potential value (R2 = 0.793), denaturation temperatures (β-conglycinin, R2 = 0.941; glycinin, R2 = 0.977), emulsifying activity index (R2 = 0.983), foaming capacity (R2 = 0.955), and trypsin inhibitor activity (R2 = 0.821) of SPI were positively correlated with phytate content, while protein content (R2 = 0.876), protein recovery (R2 = 0.781), emulsifying stability index (R2 = 0.953), foaming stability (R2 = 0.919), gel hardness (R2 = 0.893), and in vitro digestibility (R2 = 0.969) were negatively correlated with phytate content. Simulated gastrointestinal digestion and following dialysis showed that percentages of dialyzable Zn and Ca were increased with decreasing phytate content, while the amounts of dialyzable Zn and Ca revealed different behaviors: the former was increased and the latter was decreased. Circular dichroism spectra showed that secondary structure of SPI was changed by phytase. Compared with traditional processing method, phytase-assisted processing method could produce SPI with lower phytate and higher protein contents, which had better in vitro digestibility and could be used to prepare gel with higher hardness by partially losing some other functional properties.
Oil bodies (OBs), one of the major components of soymilk, are very important for the properties of soymilk and its related products. In this study, the properties and the related protein behaviors of OBs in the soymilk preparation were systematically examined. Raw soymilk OBs could keep their natural integrities after grinding, and they were bound by many soybean proteins (mainly beta-conglycinin, glycinin, and Gly m Bd 30K) that could be removed by pH 11.0 washing. Heated soymilk OBs, including heat-induced coalesced OBs (> 1 mu m) and modified OBs (around 400 nm), were strongly bound by beta-conglycinin and glycinin that even could not be removed by pH 11.0 washing. The oleosins (24 kDa, P29530 and P29531; 18 kDa, C3VHQ8) of raw soymilk OBs were hydrolyzed to 16 kDa polypeptides (on SDS-PAGE gel) when exposed to room temperature, while immediately heating, the raw soymilk could prevent the oleosin hydrolysis. Then, the isolated raw and heated soymilk OBs revealed different dispersion stabilities as a function of pH. At last, tofu curds were made from two reconstituted soymilks containing non-lipid soymilk and raw or heated soymilk OBs (pH 11.0 isolation), and one trend was found that tofu curds containing heated soymilk OBs had lower breaking stress and Young's modulus than those containing raw soymilk OBs. It is considered that this study is meaningful for designing new strategies to improve the qualities of soymilk and its related products.
In this study, soy proteins were reduced with 0.1–10 mM dithiothreitol (DTT) to obtain an increasing number of sulphydryl groups (SH) with a similar particle size. Aggregation was promoted by increasing the degree of reduction when heated (100 °C, 30 min), resulting in larger sized aggregates (from 40 to 70 nm) and a higher viscosity of the aggregate dispersion. The disulphide bond (SS) content decreased and the less SS linked polymer, which was composed of acidic (A) polypeptide of glycinin, basic (B) polypeptides of glycinin, and a small amount of α′ and α subunits of β-conglycinin, was formed with increasing reduction degree, suggesting that SH/SS polymerisation was not the driving force for aggregation. The larger aggregates with increasing degrees of reduction were composed of more B of glycinin and β of β-conglycinin, suggesting that the A and the small amount of α′ and α in the SS linked polymer have an inhibiting effect on protein aggregates formation.
Proteins in soybean whey were separated by Tricine-SDS-PAGE and identified by MALDI-TOF/TOF-MS. In addition to β-amylase, soybean agglutinin (SBA), Kunitz trypsin inhibitor (KTI), a 12 kDa band was found to have similar amino acid sequence with that of Bowman-Birk protease inhibitor (BBI) and showed both trypsin and chymotrypsin inhibitor activities. The complex behavior of soybean whey proteins (SWP) with chitosan (Ch) as a function of pH, protein to polysaccharide ratio (RSWP/Ch), was studied by turbidimetric titration and SDS-PAGE. During pH titration, the ratio of zeta potentials (absolute values) for proteins to chitosan (∣ZSWP∣/ZCh) at the initial point of phase separation (pHφ1), were equal to the reciprocal of their mass ratio (SWP/Ch), revealing that the electric neutrality conditions were fulfilled. The maximum protein recovery (32%) was obtained at RSWP/Ch =4:1 and pH 6.3, while at the RSWP/Ch =20:1 and pH 5.5, the chitosan consumption was the lowest (0.196g Ch/g recovered proteins). In the protein-chitosan complex, KTI and the 12kDa protein were higher in contents, than those of SBA and β-amylase. However, if soybean whey was pre-centrifuged to remove aggregated proteins, and interacted with chitosan at the condition of SWP/Ch=100:1, pH 4.8 and low ionic strength, KTI was found to be selectively complexed. After removing chitosan at pH 10, a high purity KTI (90% by SEC-HPLC) could be obtained.
Trypsin inhibitor activity (TIA) is an important antinutritional factor in soymilk. In this study, the effects of NaCl preaddition on TIA and the heat-induced TIA inactivation mechanisms were examined. The results showed that Kunitz trypsin inhibitor (KTI) and Bowman-Birk inhibitor (BBI) contributed 74% and 26% to raw soymilk TIA, respectively. The heat-induced quick KTI incorporation into protein aggregates was the reason for its quick TIA inactivation. The heat-induced slow cleavage of one BBI peptide bond was the reason for its slow TIA inactivation. Heat-induced protein aggregate formation had little effect on BBI inactivation owing to the fact that BBI and its degradation product tended to remain in the supernatant (197,000g, 1 h) in all conditions used in this study. NaCl could accelerate the KTI incorporation into protein aggregates and the cleavage of one BBI peptide bond, which supplied a simple and quick method for low TIA soymilk processing.
Unlabelled: In this study, the macronutrients and micronutrients of pH 6.8, 8.0, 9.5, and 11.0 extracted soybean oil bodies (OBs) were examined, revealing that soybean OBs might be used as a natural carrier for bioactive components (unsaturated fatty acids, phospholipid, tocopherol, and phytosterol). pH 6.8 extracted OBs (dry basis) contained 85.88% neutral lipid, 8.18% protein, and 5.85% polar lipid (mainly phospholipid) by gravimetric analysis. The percentage of neutral lipid was increased, while those of protein and polar lipid were decreased with increasing pH. Tocopherol (about 75 mg/100 g neutral lipid of OBs) was not affected, while phytosterol was decreased (136 to 110 mg/100 g neutral lipid of OBs) with increasing pH. The detectable total monosaccaride (galactosamine, glucosamine, and glucose) content of extracted OBs was low and also decreased (35.80 to 6.13 mg/100 g neutral lipid of OBs) with increasing pH. The protein of extracted OBs had higher percentage of essential amino acids than soybean protein isolate with tryptophan and methionine as limited amino acids. The fatty acid composition of extracted OBs was rich in linoleic acid (about 59%), oleic acid (about 20%), and α-linolenic acid (about 7%). Practical application: Oil bodies (OBs) from soybean and other plant seeds are greatly examined owing to their potential utilizations in food ingredients. The determination of its macronutrients and micronutrients would be very meaningful for its efficient utilization in the future.
Walnut milk, which is generally produced in the presence of stabilizers by homogenization, is being increasingly consumed by people in China, due to its good nutritional and functional properties. In this study, the mechanism behind the poor dispersion stability of walnut milk without stabilizer has been clarified as caused by heat-induced large aggregate formation from walnut oil bodies and protein. It was found that addition of raw soymilk into walnut milk could result in the stable mixed beverage by homogenization with subsequent heating. This was because raw soymilk played roles on decreasing the particle size of walnut oil bodies during the homogenization, and preventing the large aggregate formation from walnut oil bodies and protein during the subsequent heat treatment. Amino acid analysis showed that the stable mixed beverage contained more lysine than the walnut milk. In addition, the stable mixed beverage had higher levels of protein content (about 2%) than the commercial walnut milk (commonly < 1%) in China. Therefore, this study was meaningful for the convenient and value-added utilization of walnut.
Soybean soluble polysaccharides (SSPS) were extracted under acidic condition from the okara of producing soybean protein isolate. The methyl esterificated carboxylic group of galacturonic acid (GalA) in SSPS could be demethoxylated by alkali treatment when heated, which had effects on SSPS properties. In this study, okara was treated in six selected alkali conditions, and then SSPS were extracted under the acidic condition (115 °C, 100 min, pH 4.5). The obtained SSPS were determined to have degree of esterification (DE) of 49, 55, 60, 66, 74 and 83%, respectively. Violent alkali treatment produced SSPS with not only lower DE but also lower molecular weight (MW) compared to mild alkali treatment. MW was increased from 396 to 489 kDa with increasing DE. It was found that SSPS with lower DE possessed lower aqueous viscosity, more negative charges, better emulsifying activity in oil–in–water emulsion and stabilizing activity in acidic milk drink. When DE changed from 49 to 83%, aqueous viscosity (10% (w/w) SSPS), average particle size of fresh oil–in–water emulsion, and centrifugation precipitating rate (CPR) of acidic milk drink (pH 4.5; two-stage 30 MPa homogeneous pressure) were increased from 25.8 to 108.2 mPa•s, 1.5 to 4.0 μm, and 0.20 to 0.47%, respectively; and zeta potential (pH 7) was decreased from 17 to 10 mV. These results suggested that SSPS obtained from different alkali treatment had different functional properties.
Soy protein has been extracted and differently treated to yield products of diverse desirable characteristics. Generally, synthetic chemicals (NaOH and HCl) are used to extract the protein from n-hexane-defatted soy flour (DF). Some researchers have tried to extract it from full-fat flour (FF) using natural chemicals (NC) in response to the ever-increasing consumer preference for naturally processed foods. The effects of these chemicals on the structure of the resultant protein product are not known. This study examined oxidative and structural modification effects of NC and SC as they interact with proteins in the DF and FF systems. Significant differences (p < 0.01) were observed in all samples in lipoxygenase activity, degree of oxidation, free and total sufhydryl content and intrinsic fluorescence, which indicated increased structural changes in the FF-based samples as compared to those prepared from DF. Circular dichroism spectroscopy showed existence of α-helices and β-sheets protein secondary structures. However, DF samples showed increase in unordered structural conformation changes than the FF sample. Surface hydrophobicity and size exclusion chromatography results were more related to the extraction chemicals than the type of flour used, with samples prepared with SC showing an increase. That’s, the use of natural and synthetic protein extraction chemicals from FF and DF have different effects on the structure and degree of oxidation of the resultant proteins.
As consumer interest in natural food, in which no synthetic chemicals are used, is increasing, a traditional soy protein isolate (SPI) was prepared from full-fat flour, using amaranth (Amarathus tricolor L.) ash and lemon (Citrus limon) extracts as alternatives to NaOH and HCl, respectively. SpectrumTM descriptive analysis revealed some distinctive characteristics of the traditional SPI compared to those of conventional and commercial SPIs. Principal component analysis (PCA) results accounted for 90.11% of the samples variability on a two dimensional component space based on the attribute eigenvector loadings of ≥ 0.3, and associated the traditional SPI with opacity, pasta, sweet, sour, and bitter (principal component 1, 55.14%), but not color, cardboard, cereal, brothy, ashy, astringent, salty, and viscosity, which were also characteristic to the conventional and commercial SPIs (principal component 2, 36.53%). That’s the traditional SPI had comparatively no unique characteristic that would affect its application in food systems.
As a by-product of oil production, walnut proteins are considered as an additional source of plant protein for human food. To make full use of the protein resource, a comprehensive understanding of composition and characteristics of walnut proteins are required. Walnut proteins have been fractionated and characterized in this study. Amino acid composition, molecular weight distribution and gel electrophoresis of walnut proteins and protein fractionations were analyzed. The proteins were sequentially separated into four fractions according to their solubility. Glutelin was the main component of the protein extract. The content of glutelin, albumin, globulin and prolamin was about 72.06%, 7.54%, 15.67% and 4.73% respectively. Glutelin, albumin and globulin have a balanced content of essential amino acids, except for methionine, with respect to the FAO pattern recommended for adults. SDS-PAGE patterns of albumin, globulin and glutelin showed several polypeptides with molecular weights 14.4 to 66.2 kDa. The pattern of walnut proteins in two-dimension electrophoresis (2-DE) showed that the isoelectric point was mainly in the range of 4.8-6.8. The results of size exclusion chromatogram indicated molecular weight of the major components of walnut proteins were between 3.54 and 81.76 kDa.
The adsorption of heat-denatured soy proteins at the oil/water interface during emulsification was studied. Protein samples were prepared by heating protein solutions at concentration of 1- 5% (w/v), and were then diluted to 0.3% (w/v). The results showed that soy proteins those had been heated at higher concentrations generated smaller droplet size of emulsion. Increase in homogenizer rotating speed resulted in higher protein adsorption percentages and lower surface loads at the oil/water interface. Surface loads for both unheated and heated soy proteins were linearly correlated with the unadsorbed proteins' equilibrium concentration at varying rotating speed. With the elevation in NaCl addition level, protein adsorption percentage and surface loads of emulsions increased, while the lower droplet sizes were obtained at the ionic strength 0.1 M. The aggregates and non-aggregates displayed different adsorption behaviors when rotating speed or NaCl concentration varied.
After oil bodies (OBs) were extracted from ungerminated soybean by pH 6.8 extraction, it was found that 24 and 18 kDa oleosins were hydrolyzed in the extracted OBs, which contained many OB extrinsic proteins (i.e., lipoxygenase, β-conglycinin, γ-conglycinin, β-amylase, glycinin, Gly m Bd 30K (Bd 30K), and P34 probable thiol protease (P34)) as well as OB intrinsic proteins. In this study, some properties (specificity, optimal pH and temperature) of the proteases of 24 and 18 kDa oleosins and the oleosin hydrolysis in soybean germination were examined, and the high relationship between Bd 30K/P34 and the proteases was also discussed. The results showed (1) the proteases were OB extrinsic proteins, which had high specificity to hydrolyze 24 and 18 kDa oleosins, and cleaved the specific peptide bonds to form limited hydrolyzed products; (2) 24 and 18 kDa oleosins were not hydrolyzed in the absence of Bd 30K and P34 (or some Tricine-SDS-PAGE undetectable proteins); (3) the protease of 24 kDa oleosin had strong resistance to alkaline pH while that of 18 kDa oleosin had weak resistance to alkaline pH, and Bd 30K and P34, resolved into two spots on two-dimensional electrophoresis gel, also showed the same trend; (4) 16 kDa oleosin as well as 24 and 18 kDa oleosins were hydrolyzed in soybean germination, and Bd 30K and P34 were always contained in the extracted OBs from germinated soybean even when all oleosins were hydrolyzed; (5) the optimal temperature and pH of the proteases were respectively determined as in the ranges of 35-50 °C and pH 6.0-6.5, while 60 °C or pH 11.0 could denature them.
Understanding how foaming properties of proteins are affected by factors such as pH, salt concentration and temperature is essential in predicting their performance and utilisation. In this study, the effects of pH and salt concentration were studied on the foaming properties of pumpkin seed protein isolate (PSPI) and PSPI- xanthan (XG)/Arabic (GA) gum blends. The foaming properties of the PSPI-GA/XG blends were also compared with egg white. Foam stability (FS) was significantly affected by pH with PSPI: GA (25:4) and PSPI: XG (25:1) having a significantly higher stability at pH 2 with the lowest foam stability at pH 4. Sodium chloride (0.2-1.0 M) did not significantly affect foaming properties although PSPI: GA (25:4) had the highest FC (89.33 ± 3.24%) and FS (76.83 ± 1.53 min) at 0.2 M sodium chloride concentration. The foaming capacity (FC) of PSPI: GA (25:4) blend (128.00 ± 0.91%) was significantly higher (p < 0.05) than that of egg white (74.00 ± 1.33%) but its FS was significantly lower. It was further revealed that the FC of egg white (74.00 ± 1.33%) was comparable to the PSPI:XG (25:1) blend (74.00 ± 1.46%) but the FS for egg white (480.00 ± 2.67 min) was significantly higher (p < 0.05) than the FS (116.21 ± 0.86 min) of PSPI:XG (25:1). The foaming properties of PSPI and PSPI-xanthan (XG)/Arabic (GA) blends were significantly affected by pH. Optimum foaming properties, PSPI:XG (25:1) and PSPI:GA (25:4) were observed at pH 2 and heat treatment temperature of 80 ºC. Keywords: pumpkin seed protein isolate, xanthan gum, Arabic gum, foaming capacity, foam stability
The number of consumers preferring natural and organic foods to those that involve synthetic chemicals has recently shown dramatic growth. As such, a native isoelectrically precipitated soy protein isolates (SPIs) were prepared from full-fat and defatted flours using amaranth (Amaranthus tricolor L.) lye (pH>12.5) and lemon extract (pH<2.5) as natural food-plant-based chemicals, replacing the conventional synthetic ones (NaOH and HCl, respectively). Functional properties, total polyphenol content and antioxidant activity of the natural SPIs were compared to those of synthetic ones. All the SPI samples qualified to be soy protein isolates with a minimum protein content of 91.21%. The natural SPI showed significant increase in emulsion stability (p≤0.05). While higher values with narrow margins were shown by the synthetic than the natural SPIs in oil absorption (0.66±0.02, 0.50±0.01%, respectively), emulsion capacity (56.53±0.57, 55.50±0.39%), foam stability (11.33±0.61, 10.40±0.40%). No significant difference was observed in water absorption capacity. The DPPH assay showed increased antioxidant activity in the natural SPI although its total polyphenol content was lower. Thus, SPI with functional properties similar to those of conventional ones can be naturally processed using amaranth ash and lemon extracts as alternative chemicals to addressing
Five kinds of spacer arm attached chitosan hybrid hydrogels were tested for the possibility of being used as carriers for the immobilization of hydroperoxide lyase from amaranthus tricolor leaves. The 1,6-hexamethylenediamine attached chitosan-K-carrageenan with biomimetic hydrophobic surface was proved to be the most suitable carrier. A maximum activity of 7.49 +/- 0.19 U/g and a yield of 95% were obtained under optimized coupling condition. Meanwhile, the affinity between enzyme and substrates was not reduced after immobilization, as evidenced by the fact that the K-m value of hydroperoxide lyase decreased from 108.6 to 79.97 mu M for 13-hydroperoxy-linoleic-acid and almost unchanged for 13-hydroperoxy-linolenic-acid. Furthermore, the thermal, operational and storage stabilities of HPL were significantly improved after immobilization. Using the immobilized enzyme as the catalyst, the yield of 2(E)-hexenal and hexanal reached 1374.8 +/- 51.8 mg/L and 1987.9 +/- 67.9 mg/L, respectively, and the amount of immobilized enzyme needed in the reaction mixture was much lower than its free counterpart. Crown Copyright
Soybean oil bodies (OBs), naturally pre-emulsified soybean oil, have been examined by many researchers owing to their great potential utilizations in food, cosmetics, pharmaceutical and other applications requiring stable oil-in-water emulsions. This study was the first time to confirm that lectin, Gly m Bd 28K (Bd 28K, one soybean allergenic protein), Kunitz trypsin inhibitor (KTI) and Bowman-Birk inhibitor (BBI) were not contained in the extracted soybean OBs even by neutral pH aqueous extraction. It was clarified that the well-known Gly m Bd 30K (Bd 30K), another soybean allergenic protein, was strongly bound to soybean OBs through disulfide bond with 24 kDa oleosin. One steroleosin isoform (41 kDa) and two caleosin isoforms (27 kDa, 29 kDa), the integral bioactive proteins, were the first time to be confirmed for soybean OBs, and considerable amounts of calcium, necessary for the biological activities of caleosin, were strongly bound to OBs. Unexpectedly, it was found that 24 kDa and 18 kDa oleosins could be hydrolyzed by unknown soybean endoprotease in the extracted soybean OBs, which might give some hints for improving the enzyme-assisted aqueous extraction processing of soybean free oil.
Oscillatory tests data were analyzed using the power law and the weak gel models. Acid-induced gels had higher storage modulus (G′) than salt-induced gels. During gelation, both moduli increased gradually but never reached a steady state, with G′ being higher than G″. Under applied parameters, salt-induced gels made from soybean coagulated faster than the ones made from soy protein isolate (SPI). Syneresis rate in salt-induced gels was significantly lower than in acid-induced gels, with SPI acid-induced gel having the highest rate (36.72%) and soybean saltinduced gel having the lowest (14.77%). Furthermore, salt-induced gels resulted in a reduction in shear deformation at fracture and an increase in the shear stress at yielding, whereas the shear stress at yielding was reduced and the shear deformation at fracture was increased in both acid-induced gels. It was observed that gels made from soybean had a more uniform and denser network with smaller network pore size than the ones made from SPI.
Hydroperoxide lyase is the key enzyme in lipoxygenase pathway producing green-note flavours and has potential value for the flavour additive industry. So far, only a low yield of green-note flavours produced by hydroperoxide lyase has been achieved, primarily because of its instability. The aim of this study was to stabilise hydroperoxide lyase from Amaranthus tricolor leaves by immobilisation and investigate the characteristics of immobilised enzyme in comparison with free and native membrane-bound enzyme. A maximum activity of 2.85 ± 0.1 U g−1 (wet) ceramic hydroxyapatite and a yield of 80% were obtained under optimised coupling conditions. The optimal reaction pH was 6.0, 6.0 and 7.5 for free, membrane-bound and immobilised enzyme respectively, while the optimal reaction temperature was 30, 35 and 35 °C respectively. Thermal and operational stability of immobilised enzyme were substantially enhanced. However, a higher substrate diffusion resistance was imposed after immobilisation, as evidenced by the Km value of immobilised enzyme being higher than that of free and membrane-bound enzyme. Ceramic hydroxyapatite was a candidate for the immobilisation of hydroperoxide lyase from A. tricolor leaves. The stability of hydroperoxide lyase was substantially improved after immobilisation on this substrate.
Mixtures of soy protein (SP) and gum arabic (GA) formed an electrostatic complex in a relatively narrow pH range at very low ionic strength. The conditions under which the complexes could be formed were determined using turbidimetric measurements first. In salt-free condition and 1:1 SP/GA mixture, critical pH values with the formation of soluble (pHc = 4.40), insoluble (pHφ1 = 3.55), and maximum (pHopt = 3.15) complexes were observed. As SP/GA ratios increased from 1:4 to 8:1, critical pH values shifted toward higher pH. Charge densities (ZN) for SP and GA were calculated from electrophoretic mobility using soft particle analysis theory. Results showed that a 1:1 charge ratio at pHφ1 was found at any SP/GA ratio, indicating that charge compensation was fulfilled for SP/GA insoluble complex formation. A SP-GA-water ternary phase diagram was built at pH 4.0. The influence of the total biopolymer concentration (0-10% w/w) and SP/GA ratio was represented in the phase diagram. At a total concentration of 0.10%, results were consistent with the turbidity measurement; that is, no phase separation occurred at an SP/GA ratio lower than 1:2 at pH 4.0. Salt effect (NaCl, 0-500 mmol/L) on SP/GA complexes was discussed. Results indicated that SP/GA complexing, which led to the formation of turbidity peaks at pH 3.2, was suppressed when NaCl concentrations were ≥50 mmol/L, whereas the remarkable increase in turbidity around pH 5.0 was caused by the aggregation of soy protein molecules on which gum arabic could be adsorbed.
The fermented sorghum flour paste (ajena), is one of the most common traditional cereal based food products in Africa, especially in Sudan and Ethiopia. It is used as a main ingredient or starting material for other food products. This traditional fermentation biotechnology has recently caught the attention of food scientists trying to improve various properties of the product. In this study, the functional and protein quality properties of naturally fermented blended flour made up of 20% native broad bean and 80% sorghum flours were compared with conventionally fermented sorghum flour. The blended flour increased in protein composition by about 5% from 11.46 to 16.37 after fermentation. The protein quality in terms of in vitro protein digestibility (IVPD) increased from 58.53 % in fermented sorghum flour to 71.93 % in the blended flour. The increased protein, improved (p≤0.05) the functionality of the blended flour as reflected in its better water-absorption capacity, compared to value obtained from fermented sorghum for this functional property. Bulk density and swelling power of the two flours showed no difference. The Mixolab results indicated lower dough stability, elasticity and consistency in the blended flour as compared to non-blended sorghum flour samples. Thus, ajena, with improved nutrition in terms of protein and less elasticity, like those used in cereal based drinks can be processed by blending sorghum and broad bean flours before fermentation.
The storage modulus (G′) and fracture properties of the non-treated and NEM-treated emulsion gels were investigated in the absence and presence of unadsorbed soy protein aggregates (USPA). In the absence of USPA, a decrease in the G′ of emulsion gels was observed after NEM treatment. However, in the presence of USPA, the addition of NEM only slightly decreased the G′ (p < 0.05). For both non-treated and NEM-treated emulsions, a ∼63-folds higher G′ value was obtained after the USPA addition. These results revealed the presence of sulphydryl group – disulfide bond interchange reactions at ambient temperature and under acidic conditions. In the absence of USPA, the sulphydryl group – disulfide bond interchange reactions was the main interactions responsible for the higher G′ values of non-treated emulsion gels, but for the emulsions with USPA presented, the large quantity of non-covalent interactions (e.g. hydrophobic group & hydrogen bonds) is the main interactions responsible for the aggregation and gelation of emulsion droplets. In the presence of USPA, the sulphydryl group – disulfide bond interchange reactions formed in the non-treated emulsion gels did not increase the final G′ but increased the stability of network. A power law relation was observed between the USPA concentration and the final G′, as well as between the oil volume fraction and the fracture stress/strain.
In this study, the sulfhydryl (SH) contents of unheated and heated (90°C, 5 min) soy protein were detected under different conditions (pH, reagent addition order, SDS/GuHCl concentration, EDTA) using two aromatic disulfide reagents: 5, 5'-dithiobis-(2-nitrobenzoic acid) (DTNB) and 4, 4'-dithiodipyridine (DPS). Two fluorescent alkylating reagents, monobromobimane (mBBr) and N-(1-pyrenyl)-maleimide (NPM), were chosen due to their high sensitivity and were also used. Amino acid analysis was used to detect the SH (cysteine) contents of unheated (7.51 ± 0.45 μmol SH/g protein) and heated (1.47 ± 0.10 μmol SH/g protein) soy protein, and similar results were obtained using enzymatic hydrolysis-assisted DPS. The SH content detected by DTNB was affected by pH, denaturant species, and denaturant concentration, and the best results were obtained at pH 7.0 when 6 M GuHCl was added after DTNB. These results were lower than that of the amino acid analysis, however. The SH detected by DPS was not as affected as that of DTNB by pH, denaturant species, and denaturant concentration. Additionally, the results of the amino acid analysis were similar to that of DPS at pH 7.0 in 2% SDS and 4 to 6 M GuHCl when SDS and GuHCl were added after DPS. EDTA did not have a significant effect on SH detection when DTNB and DPS were added before SDS and GuHCl. Finally, although mBBr and NPM can detect SH in low protein concentrations (1/10 of that required for DTNB and DPS), mBBr and NPM overestimated the SH content of soy protein. Therefore, using DPS at pH 7.0 when it is added before SDS and GuHCl is the most reliable method for detecting the SH content of soy protein.
With increasing preference for all-natural foods to those involving synthetic chemicals, native isoelectrically precipitated soy protein isolate (SPI) was prepared using amaranth (Amaranthus tricolor L.) lye (pH > 12.5) and lemon extract, (pH < 2.5) as natural, food-plant-based chemicals. Protein content (91.21 %), yield (43.62 %) and digestibility correlation amino acid score (0.77) were obtained and were comparable to those of SPI prepared using synthetic chemicals (NaOH and HCl). Methionine and cystein-s were significantly higher in the natural SPI while glutamine and serine were higher in synthetic SPI (p < 0.01). Most of the determined minerals were higher in the natural SPI with potassium being the highest. Sodium was very high in the synthetic SPI. The rest of the minerals including phosphorus, iron and nickel, showed no significant difference. Anti-nutritional factors (trypsin inhibitors and phytic acid) were considerably lower in the natural SPI. Thus, a quality all-natural SPI can be produced using amaranth lye and lemon extract to address concerns regarding use of synthetic chemicals.
Adsorption behavior of daidzin, genistin, 6″-O-malonyldaidzin and 6″-O-malonylgenistin, the four major soy isoflavones presented in soy molasses centrifugation supernatant on activated carbon was studied in this paper so as to provide theoretical basis for the purification of soy oligosaccharides from soy molasses. Kinetic experiments showed that the adsorption processes obeyed pseudo-second-order kinetics and equilibrium was nearly achieved in 90 min. Weber–Morris model fitting showed that adsorption process consisted of 3 stages: boundary layer diffusion and two intra-particle diffusions. Experimental adsorption data for every isoflavone components could be described separately by the Langmuir isotherm model and the calculated maximum adsorptions were in the order of genistin > daidzin > 6″-O-malonylgenistin > 6″-O-malonyldaidzin, indicating that the adsorption driving forces were due to dispersion interactions between the aromatic ring of isoflavone and the aromatic structure of the activated carbon. Adsorption behaviors of isoflavones on activated carbon in sugar free solutions were compared. It was found that, by removing sugar from the system, diffusion rate constants and the sum of the maximum adsorption capacity increased.
Oxidative modification of soy protein by lipid peroxidation products, which was potentially present in a lipoxygenase-catalyzed polyunsaturated fatty acid peroxidation system, was investigated in this study. 13S-Hydroperoxy-9Z, 11E-Octadecadienoic acid (HPODE), malondialdehyde and acrolein were selected as representative primary product and secondary byproducts of lipid peroxidation and 2, 2'-azobis-(2-amidinopropane) dihydrochloride (AAPH) -derived peroxyl radical peroxyl radicals were chosen to simulate lipid peroxidation-derived free radical. Incubation of soy protein with increasing concentration of AAPH, HPODE, malondialdehyde and acrolein resulted in gradual generation of protein carbonyl derivatives, loss of free sulphydryl groups, total sulphydryl groups, free amine, available lysine, surface hydrophobicity and formation of oxidation aggregates. The average distribution model of protein accessible groups could explain majority mechanism of lipid peroxidation products-mediated soy protein oxidation. Primary oxidation aggregates further developed into insoluble aggregates by covalent cross-linking also may provide a partial mechanism of lipid peroxidation products-mediated soy protein oxidation.
In this study, composition, structure and the functional properties of protein concentrate (WPC) and protein isolate (WPI) produced from defatted walnut flour (DFWF) were investigated. The results showed that the composition and structure of walnut protein concentrate (WPC) and walnut protein isolate (WPI) were significantly different. The molecular weight distribution of WPI was uniform and the protein composition of DFWF and WPC was complex with the protein aggregation. H(0) of WPC was significantly higher (p < 0.05) than those of DFWF and WPI, whilst WPI had a higher H(0) compared to DFWF. The secondary structure of WPI was similar to WPC. WPI showed big flaky plate like structures; whereas WPC appeared as a small flaky and more compact structure. The most functional properties of WPI were better than WPC. In comparing most functional properties of WPI and WPC with soybean protein concentrate and isolate, WPI and WPC showed higher fat absorption capacity (FAC). Emulsifying properties and foam properties of WPC and WPI in alkaline pH were comparable with that of soybean protein concentrate and isolate. Walnut protein concentrates and isolates can be considered as potential functional food ingredients.
The protein concentration dependence on the rheological properties of acid-induced gels formed with unheated and heated soy protein-stabilized emulsions (UHSPE and HSPE) was investigated at different acidification temperatures. Pre-heat treatment on soy protein solutions resulted in a higher storage modulus (G′) and a shorter gelation time (tgel) of acid-induced emulsion gels. A maximum in tan δ was observed in the UHSPE gels but no maximum was detected in the HSPE gels. Increasing the acidification temperature decreased the G′ and tgel. The dependence of the G′ on the protein concentration (c) can be scaled with a power law: G′ ∼ cA. The exponent (A) increased with pre-heat treatment and acidification temperature. The experimental data.fitted the fractal scaling model (G′ ∼ φA) and the simple time- scaling model above very well for the acid-induced soy protein-stabilized HSPE gels with varying oil volume fraction. The large deformation and fracture properties were significantly affected by soy protein concentration, pre-heat treatment, acidification temperature and volume fraction of oil droplets (p < 0.05).
Defatted soybean flour was treated with hexane and ethanol to reduce lipid content and heated to inactivate lipoxygenase (LOX, linoleate:oxygen reductase; EC 22.214.171.124) to obtain lipid-reduced soybean flour (LRSF). The effects of processing conditions such as pH, reducing agent and storage time on yields and purity of glycinin (11S) were evaluated in the fractionation of soybean glycinin isolated from LRSF. Adjusting the pH of protein extract from 6.2 to 6.6, the yield of glycinin decreased by 16.71%, while the purity of the protein increased by 4.60%. Sulfhydryl and disulfide content of proteins increased by degrees with increasing pH. Compared with dithiothreitol (DTT) or β-mercaptoethanol (ME) as reducing agent, the yield of glycinin was the highest when sodium bisulfite (SBS) was added to the protein extract at pH 6.4. The effect of DTT on yields of glycinin was the lowest of the three kinds of reducing agent. The purity of glycinin was similar when the three kinds of reducing agent were used. These results showed that SBS was the best choice for the isolation of 11S-rich fraction. Prolonging storage time in the precipitation stage, 10 h was the best for yields and purity of glycinin in the experiment, while there was no significant difference at P ≥ 0.05 for total sulfhydryl and disulfide content. The decreased free sulfhydryl content of glycinin indicated that the oxidation of free sulfhydryls and the formation of disulfide bonds occurred when the extraction time was prolonged.
(2E)-Hexenal is widely used in flavors and perfumes; however, it is mainly derived from chemical synthesis. In this study, hydroperoxide lyase (HPL) naturally originated from Amaranthus tricolor was used to catalyze the reaction to generate (2E)-hexenal from 13-hydroperoxy-9Z, 11E, 15 Z-octadecatrienoic acid (13-HPOT), and salt-adding steam distillation was used for the separation of (2E)-hexenal. A maximum yield of (2E)-hexenal that reached 1,156.4 mg L−1 was obtained with a high substrate concentration (40 mM 13-HPOT) under the following conditions: 10 min, pH 7.5, 20 °C, HPL 16 U mL−1, butylated hydroxytoluene (BHT) 1 mM, and dithiothreitol (DTT) 15 mM. Then the separation of (2E)-hexenal was conducted by salt-adding steam distillation. It was found that AlCl3 had the greatest effect on the separation of (2E)-hexenal, followed by CaCl2 and NaCl. The distillate yield with the addition of AlCl3 was 93.2 %, while the distillate yield without the addition of salt was 81.2 %. The distillate concentration with AlCl3 was 6,464.2 mg L−1, while the distillate concentration without the addition of salt was 4,490.3 mg L−1. The addition of salt improved the efficiency of the steam distillation. This study was greatly meaningful for providing a green method to the large-scale production of (2E)-hexenal.
Kunitz trypsin inhibitor (KTI) and Bowman-Birk inhibitor (BBI) have trypsin inhibitor activities (TIA), which could cause pancreatic disease if at a high level. It is not clear why some KTI and BBI lose TIA and some does not in the soymilk processing. This would be examined in this study. TIA assay showed residual TIA was decreased with elevated temperature and TIA was decreased quickly in the beginning and then slowly in boiling water bath. Interestingly, ultracentrifugation showed low residual TIA soymilk had more precipitate than high residual TIA soymilk and soymilk TIA loss had a high correlation coefficient (R(2) > 0.9) with precipitate amount. In addition, the TIAs of floating, supernatant, and precipitate obtained by ultracentrifugation were assayed and >80% residual TIA was concentrated in the supernatant. Tricine-SDS-PAGE showed KTI in supernatant was mainly a noncovalent bound form which might exist as itself and/or incorporated into a small protein aggregate, while KTI in precipitate was incorporated into a protein aggregate by disulfide and/or noncovalent bonds. Chymotrypsin inhibitor activity (CIA) assay showed about 89% of the original CIA remained after 100 °C for 15 min. Ultracentrifugation showed that >90% residual CIA was concentrated in supernatant. Tricine-SDS-PAGE showed soymilk (100 °C, 15 min) BBI mainly existed in supernatant but not in precipitate. It was considered that BBI tended to exist as itself with its natural conformation. Thus, it was suggested residual TIA was mainly from the free BBI and TIA inactivation was mainly from KTI incorporation into protein aggregate. This study is meaningful for a new strategy for low TIA soymilk manufacture based on the consideration of promoting protein aggregate formation.
A cDNA encoding hydroperoxide lyase (HPL) was isolated from Solanum tuberosum, cloned into pQE-30 vector, and expressed in E. coli. The recombinant protein was purified by nickel affinity chromatography and showed an approximate molecular weight of 54 kDa by SDS–PAGE analysis, which was similar to the predicted value based on the putative amino acid sequences (53.9 kDa). 13-Hydroperoxy-linolenic acid (13-HPOT) was the preferred substrate for the enzyme compared with 13-hydroperoxy-linoleic acid (13-HPOD). The corresponding volatile products were 2(E)-hexenal and n-hexanal tested by headspace-gas chromatography, respectively. The enzyme was optimally active at 25 °C and pH 6.5. The K m, V max, and the catalytic efficiency (V max/K m) for 13-HPOT were 56.6 μM, 71.3 units/mg, and 1.26 units/mg · μM, respectively. Activity of the recombinant potato HPL increased when Triton X-100, sodium chloride, or potassium chloride was added in the reaction mixture, while calcium chloride decreased activity of the recombinant enzyme.
A novel, rapid and ultrasensitive surface-enhanced Raman scattering (SERS) immunoassay for the detection ofmicrocystin LR was developed based on the assembly of goldnanorods (GNRs). GNRs were assembled into nanorod chains through the bio-recognition. The SERS signal of the probe molecule modified to the end of the NRs could be enhanced due to the hot spot between the NRs. Meanwhile, the finite integration technique simulation revealed that the electrical field of nanorods was increased obviously depending on the degree of end to end assembled structures. The research results demonstrated that the linear detection range was from 0.01 ng mL−1 to 5 ng mL−1, the limit of detection was 5 pg mL−1, and the time necessary for the analysis was only 15 min.
Sorghum germination resulted in a substantial tannin loss (95.7 %). Proximate composition, titratable acidity, pasting properties, in vitro protein digestibility, and protein solubility were studied post fermentation (Saccharomyces Cerevisiae) of the blended soybean, maize, and germinated sorghum flours. The pH progressively decreased with fermentation time, while titratable acidity increased from 0.029 to 0.118 ml/ml. Crude protein content increased with fermentation (251.7-274.8 mg/g) as a result of a shift in the dry matter composition. In-vitro protein digestibility markedly increased (12 %) as a result of fermentation. Protein solubility curves were above 30% of which highest for both fermented (12 and 24 hours) and unfermented composite flours were at pH 12 (51.77-77.64%) and lowest at pH 4 (30.31-35.98%). SDS-PAGE showed that protein hydrolysis occurred during fermentation over 12 and 24 hours. Unfermented composite flour was potentially stable as food ingredient due to its pasting stability, but the fermented flour low viscosity potential was preferred in this study as more flour will be used during porridge making, hence giving a food with a high nutrient density.
The low solubility of wheat gluten is one of the major limitations to its use in food processing, and enzymatic hydrolysis has been found to be an effective way to prepare more soluble bioactive peptides from gluten. The aim of this study was to prepare bioactive peptides from modified wheat gluten (MWG) in a continuous enzymatic membrane reactor (EMR) that allowed rapid separation of low-molecular-weight peptides from hydrolysates, thus avoiding the disadvantages of batch reaction such as inefficient use of enzymes, inconsistent products due to batch-to-batch variation, substrate-product inhibition, low productivity and excessive hydrolysis. Wheat gluten was modified to decrease its lipid and starch contents in order to prevent membrane fouling. The optimal working conditions for Alcalase to hydrolyse MWG in the EMR were a substrate concentration of 20 g L(-1) , an enzyme/substrate ratio of 0.03, an operating pressure of 0.04 MPa, a temperature of 40 °C and a pH of 9. The operating stability of the EMR (including residual enzyme activity, productivity and capacity) was high. The permeate fractions showed antioxidant activities that were mostly due to low-molecular-weight peptides. A simple theoretical kinetic model was successfully applied to the enzymatic hydrolysis of MWG in the EMR. Modification of wheat gluten made the continuous enzymatic membrane reaction more efficient and the EMR proved to be an effective means of producing peptides with particular properties and bioactivities. The permeate fractions (mainly < 1000 Da) were homogeneous and stable and also showed strong antioxidant activities.
Soy protein, an important efficient emulsifier, is widely used by the food industry for incorporation into milk, yogurts, ice cream, salad dressings, dessert products, etc. The objective of this study was to investigate the rheological and physical properties of soy protein-stabilised emulsion gels as affected by protein concentration and gelation temperature. The rheological properties and permeability were determined using oscillatory rheometry, permeability and whey separation. The modulus (G' and G″), fracture stress and fracture strain of acid-induced emulsion gels after 20 h of glucono-δ-lactone addition depended strongly on soy protein concentration and gelation temperature. At increasing soy protein concentrations, acid-induced emulsion gels had shorter gelation times but higher storage moduli (G'), fracture stresses and strains. Increasing gelation temperature decreased the gelation time, G', fracture stresses and strains. Permeability and whey separation were significantly affected by the protein concentration and the gelation temperature. A significant positive correlation was observed between whey separation and permeability coefficient in emulsion gels formed at different temperatures. The rheological properties and permeability of soy protein-stabilised emulsion gels were significantly influenced by protein concentration and gelation temperature.
In this work, we report on the application of versatile gold nanorods (GNRs) in optical sensors for the detection of antibiotics. The target analyte, Gentamicin (GM) and ovalbumin (OVA)-antigen-modified GNRs together competed with antibody-modified GNRs, then influenced the formation of side-by-side aggregates of the GNRs by antibody-antigen interactions. Accordingly, the UV-vis absorption intensity of the side-by-side aggregates was changed in the presence of the target analyte. This assay allowed the selective determination of GM in the range of 0.1-20 ng/mL, and the limit of detection (LOD) of GM was 0.05 ng/mL. Furthermore, compared with the traditional plate-based immunoassay, the developed method was easy to perform without washing cycles and the results could be read as soon as the nanoprobe-analyte incubation was complete. Therefore, the developed method could be a promising tool for the detection of antibiotic residues.
Effects of protein oxidation on thermal aggregation and gel properties of soy protein by 2,2′-azobis (2-amidinopropane) dihydrochloride (AAPH)-derived peroxyl radicals were investigated in this article. Incubation of soy protein to increase concentration of AAPH resulted in a decrease in particle size and content of thermal aggregates during thermal-induced denaturation. Protein oxidation resulted in a decrease in water-holding capacity (WHC), gel hardness and gel strength of soy protein gel. An increase in coarseness and interstice of the gel network was accompanied by uneven distribution of interstice as extent of oxidation of soy protein increased. A decrease in disulphide content and formation of oxidation aggregates in the process of oxidative modification were contributed to the decline of particle size and content of thermal aggregates during thermal-induced denaturation, leading to a decrease in WHC, gel hardness and gel strength of soy protein gel.
The effect of heat treatment on the properties of soy protein-stabilised emulsions was investigated. Emulsions were prepared with unheated and heat-treated soy protein (NSP and HSP) dispersions. Heating on soy protein dispersions at 95 °C for 30 min resulted in smaller average oil droplet size, lower tendency for oil droplet flocculation, higher protein adsorption and lower viscosity. The properties of emulsions were significantly influenced by the protein concentration. The sodium dodecyl sulphate–polyacrylamide gel electrophoresis (SDS–PAGE) profiles showed that the heat treatment on soy protein dispersions increased the protein adsorption at O/W interface. The viscosity of all samples at low shear rate was inversely proportional to the d32, suggesting a positive relation to the total interfacial area per unit volume. Emulsions showed shear-thinning behaviour. The relaxation time was found to increase with aqueous phase viscosity determined by the Cross viscosity model.
Hydroperoxide lyase (HPL) was extracted from amaranth tricolor leaves using Triton X-100, and purified to electrophoretic homogeneity by ammonium sulfate precipitation, ion-exchange chromatography, hydrophobic interaction chromatography and hydroxyapatite chromatography. The purified HPL preparation consisted of a single band and spot with a molecular mass of about 55kDa as shown in SDS–PAGE and 2-D PAGE, respectively; the isoelectric point was found to be about 5.4. The maximum activity of the enzyme was observed at pH 6.0 and 25°C, respectively. The HPL showed higher activity against 13-hydroperoxy-linolenic acid compared to 13-hydroperoxy-linoleic acid. K m value for 13-hydroperoxy-linolenic acid was 62.7μM, and the corresponding V max was 178.5μMmin−1. The activity of HPL was significantly inhibited by nordihydroguaiaretic acid, HgCl2 and 2(E)-hexenal but not by EDTA and hexanal. KeywordsAmaranth tricolor-C6 aldehydes-Hydroperoxide lyase-Purification
Hydroperoxide lyase (HPL) has potential value for the flavour additive industry. Currently, the production and application of HPL suffer from stability problems. The objective of this study was to investigate the stabilisation of HPL preparation from Amaranthus tricolor leaves by the addition of selected chemical additives. Amaranthus tricolor leaves were identified as a particularly rich source of 13-HPL activity. The addition of 100 g L(-1) sucrose and trehalose to microsomal HPL prior to lyophilisation could retain nearly 100% enzymatic activity, compared to only 20% for the lyophilised control. The lyophilised microsomal HPL containing sucrose maintained full activity for even 40 days storage at -20 degrees C. For HPL solution, glycerol was effective for long-term stability at -20 degrees C. Moreover, poyols (sucrose and trehalose) and amino acid (glycine) enhanced the thermostability of HPL, while KCl and polyol mannitol decreased the thermostability of HPL. The flavour-producing enzyme HPL, found in the leaves of Amaranthus tricolor, was stabilised by the addition of chemical additives.
Microstructure and rheological properties of mixtures of acid-deamidated rice protein (ADRP) and dextran were studied. The microstructures of the ADRP/dextran mixtures were described using confocal laser scanning microscopy (CLSM), which revealed the effective association among ADRP and formation of a protein network-like structure in the mixture with higher protein concentration. Mechanical properties of the mixtures were observed by rheometer. The steady shear measurements showed a correlation with the CLSM results via a marked increase in the viscosity of the mixtures with protein association. Frequency sweeps further evidenced the build-up of the gelled network-like structure. The differences in fracture forces observed by textural measurements between mixture and single ADRP gel also suggested the difference in microstructures. The formation of network-like structure appeared to have occurred through a phase separation of ADRP and dextran.
Acrolein was selected as a representative secondary byproduct of lipid peroxidation to investigate the effect of oxidative modification of reactive aldehyde on soy protein structure. Acrolein reacted with histidine, lysine and cysteine residues in soy protein to form covalent adducts, leading to protein carbonylation and degradation of sulfhydryl groups. Circular dichroism spectra showed that soy protein modification by acrolein was related to loss of α-helix and increase of β-sheet structure. The decrease in solubility, surface hydrophobicity and intrinsic fluorescence indirectly implied that acrolein induced soy protein aggregation, and results obtained by size-exclusion chromatography directly showed that gradual aggregation of soy protein was induced by increasing concentration of acrolein. Results of sodium dodecyl sulfate polyacrylamide gel electrophoresis indicated that acrolein caused soy protein cross-linking which non-disulphide covalent bonds were involved in the formation of cross-linking, and subunits of β-conglycinin were more vulnerable to acrolein than that of glycinin.
Oxidative modification of soy protein by peroxyl radicals generated in a solution containing 2,2’-azobis (2-amidinopropane) dihydrochloride (AAPH) under aerobic condition was investigated. Incubation of soy protein with increasing concentration of AAPH resulted in gradual generation of protein carbonyl derivatives and loss of protein sulphydryl groups. Circular dichroism spectra indicated that exposure of soy protein to AAPH led to loss of α-helix structure. Effect of oxidation on tertiary structure was demonstrated by surface hydrophobicity and tryptophan fluorescence. Surface hydrophobicity steadily decreased, accompanied by loss and burial of some tryptophan residues, indicating that soy protein gradually aggregated. The results of the size exclusion chromatogram (SEC) implied that incubation caused an AAPH-dose-dependent increase of fragmentation and aggregation of oxidised soy protein. Sodium dodecyl sulphate–polyacrylamide gel electrophoresis (SDS–PAGE) indicated that non-disulphide linkages were involved in aggregate formation, and β-conglycinin was more vulnerable to peroxyl radicals than glycinin.
To reveal the role of primary products of lipid peroxidation during soy protein oxidation process, oxidative modification of soy protein by 13-hydroperoxyoctadecadienoic acid (13-HPODE) generated by lipoxygenase-catalyzed oxidation of linoleic acid was investigated in this article. Incubation of soy protein with increasing concentration of 13-HPODE resulted in generation of protein carbonyl derivatives and loss of protein sulfhydryl groups. Circular dichroism spectra indicated that exposure of soy protein to 13-HPODE led to loss of α-helix structure. Effect of oxidation on tertiary structure was demonstrated by surface hydrophobicity and tryptophan fluorescence. Surface hydrophobicity gradually decreased, accompanied by loss and burial of some tryptophan residues. The results of surface hydrophobicity and tryptophan fluorescence implied that aggregation was induced by oxidation. Size exclusion chromatogram indicated that the extent of aggregation was increased in a 13-HPODE dose-dependent manner. Sodium dodecyl sulfate polyacrylamide gel electrophoresis indicated that non-disulfide linkages were involved in aggregate formation, and β-conglycinin was more vulnerable to 13-HPODE than glycinin.
BACKGROUND: Protein oxidation results in covalent modification of structure and deterioration of functional properties of target protein. Oxidation extent of soy protein was affected by the content and type of lipid peroxidation (LPO) products in defatted soybean flours during storage and processing. Malondialdehyde (MDA) was selected as a secondary byproduct of LPO to investigate the effects of oxidative modification of LPO-derived reactive aldehyde on soy protein structure.RESULTS: MDA reacted with ε-amino and sulfhydryl groups of soy protein, and resulted in an increase in protein carbonyl groups but a decrease in sulfhydryl/disulfide, free amines and lysine. The decrease in solubility, surface hydrophobicity and intrinsic fluorescence indirectly indicated that MDA induced soy protein aggregation, and results of high-performance size-exclusion chromatography directly showed that gradual aggregation of soy protein was induced by increasing concentration of MDA. Results of electrophoresis indicated that MDA caused soy protein aggregation, and non-disulfide covalent bonds were involved in aggregate formation.CONCLUSION: The results showed that sensitivity of soy protein was related to MDA concentration. Soy protein gradually aggregated with increase of MDA concentration; β-conglycinin was more sensitive to MDA modification than glycinin. Copyright © 2009 Society of Chemical Industry
The effects of ionic strength on heat-induced aggregation of soy protein and phase separation of different soy protein aggregates with dextran were investigated. The increase of ionic strength accelerated protein aggregation as shown by an increase in turbidity, aggregate fraction and particle size of salt-induced aggregates (SA). Adding salt (NaCl) to the aggregates formed at the ionic strength of zero (non-salt aggregates, non-SA), the increase of aggregate size was also found. Zeta potential results evidenced the charge screening effects of NaCl. The results of phase diagrams indicated that the compatibility of mixtures at higher ionic strength was lower than those at lower ionic strength, and SA was more incompatible with dextran than non-SA. The effects of the increase of aggregate size on the phase separation outweighed the ionic strength, which indicated that the depletion interaction also played an important role in the phase separation of soy protein aggregates and dextran. CLSM (Confocal Laser Scanning Microscopy) and rheological observations provided additional information of the microstructures of the mixtures.
Soy protein hydrolysates with lower molecular weight were enzymatically prepared by several commercially available proteases (Alcalase 2.4L, Flavourzyme, Trypsin, Papain, Protease A and Peptidase R) with protein recovery varied from 42.59% to 79.87%. Relative content of positively charged peptides was determined on SP Sephadex C-25 using gradient sodium chloride solution as eluents. Immunomodulating properties were evaluated by measuring their effect on in vitro proliferation of murine spleen lymphocytes and phagocytic activity of peritoneal macrophages. The results showed that soy protein hydrolysates (SPHs) prepared with Alcalase and insoluble soy protein (InSP), preferable to other enzymes and soy proteins, have the highest immunomodulating activity and the optimum conditions were determined as follows: E/S=2% (Alcalase), 60 degrees C, pH 8.0, InSP concentration 6% and 225min. Positive correlations were obtained between the immunomodulating activity and content of positively charged peptides. The results suggested that lower molecular weight and positively charged peptides released from soy protein were effective in stimulating immunomodulating activity, thus provided insights into the preparation of potent immunomodulating products.
Phase behavior and microstructure of preheated soy proteins (heat-induced aggregates)/κ-carrageenan (κ-car) mixtures have not been reported before. Hereby, heat-induced aggregates of different sizes and κ-car mixed systems were investigated in 0.1 M NaCl with pH 7.0 at room temperature (25 °C) with native soy protein/κ-car system as the comparison. Phase diagrams were established by centrifugation, chemical assays and visual observation and it was found that the larger aggregate/κ-car system had narrower stable region. The microstructures of the phase-separated mixtures were described using confocal laser scanning microscopy (CLSM), which revealed the association of protein aggregate structures after phase separation. The image analysis based on the CLSM images of different mixtures showed that the variance and the histogram of the gray values were different significantly. In addition, information of kinetics of phase separation was obtained from small deformation rheology (G′, G″).
The opioid activity of wheat gluten hydrolysates (WGHs) prepared with several proteases was investigated in vitro by the contraction of guinea pig ileum (GPI), and WGHs with high opioid activity were further obtained through desalting and ultrafiltration (UF). The opioid activity varied with different wheat gluten hydrolysates. AWGH (WGH prepared with Alcalase), PPWGH (WGH prepared with Pepsin followed by Pancreatin) and PWGH (WGH prepared with pepsin) had relatively stronger inhibitory effects on the contraction of GPI with IC50 values of 1.21±0.25, 1.29±0.38 and 1.57±0.21mg protein/ml, respectively. Subsequently, AWGH and PPWGH were desalted using cation exchange resin and anion exchange resin, with high nitrogen recovery (88.51 and 89.28%, respectively) and high desalting ratio (84.67 and 85.20%). The resultant hydrolysates were further fractionated by UF performed with a 3kDa molecular weight cut-off membrane. Compared with AWGH and PPWGH, the 3kDa-permeates have higher opioid activities with high protein content above 90% and low ash content about 1.5%. The molecular weight distributions of the two 3kDa-permeates were concentrated below 2,000Da.
BACKGROUD: Many hydrolysates from animal and plant proteins have been found to possess physiological activities. Wheat gluten, an important by-product of the wheat starch industry, is produced worldwide in enormous quantities. In this study, wheat gluten hydrolysates (WGHs) were obtained by enzymatic hydrolysis and fractionated using ultrafiltration membranes. The antioxidant activities of the hydrolysates were investigated by various antioxidant assays, including the ability to inhibit the autoxidation of linoleic acid and the scavenging effect on free radicals. Amino acid composition and molecular weight distribution were also evaluated to determine their relationship with antioxidant activity.RESULTS: The pepsin hydrolysate (PeWGH) had the highest activity and was ultrafiltrated into three major types, PeWGH I (5–10 kDa), PeWGH II (3–5 kDa) and PeWGH III (<3 kDa). PeWGH III showed stronger inhibition of the autoxidation of linoleic acid and higher scavenging activity against 2,2-diphenyl-1-picrylhydrazyl, superoxide and hydroxyl free radicals. Furthermore, PeWGH III had the highest total hydrophobic amino acid content (45.11 g per 100 g protein), and its molecular weight distribution ranged from 1700 to 100 Da.CONCLUSION: The low molecular weight and amino acid composition of PeWGHs were found to be strongly correlated with their antioxidant activity. PeWGH could be used as a natural antioxidant in the pharmaceutical and food industries in the future. Copyright © 2008 Society of Chemical Industry
The effects of molecular weight of dextran on the phase behavior and microstructure of preheated soy protein (heat-induced soy protein aggregate)/dextran mixtures have not been reported before. Hereby mixed systems of heat-induced aggregates with different size and dextran with different molecular weight have been investigated at room temperature (25 °C) and pH 7.0. Phase diagrams were established by centrifugation, chemical assays and visual observation. The mixture of dextran with larger molecular weight and larger aggregate phase separated at lower biopolymer concentration. The microstructures of the phase separated mixtures were described using confocal laser scanning microscopy (CLSM), which revealed the association of protein aggregates. The image analysis of the CLSM images showed that the histograms of the gray values were different significantly. Observations of small deformation rheology (G′, G′′) of the mixed system at concentrations corresponding to those of CLSM measurements provided additional information of the microstructures.
Soybean lipoxygenase was inactivated to different degrees by dry heating of defatted soybean flour for 0, 5, 10, 15, 20 and 25 min and soy protein isolates were prepared thereof by isoelectric precipitation of the water extract of the defatted soybean flour. The fluorescence emission intensity at 420 nm of the chloroform–methanol extract of soy protein isolates, which was indicator of the existence of peroxidized lipid, varied in parallel with the lipoxygenase residual activity in defatted soybean flours. The dispersion of soy protein isolate showed an increasing turbidity with the increase of lipoxygenase residual activity in the starting defatted soybean flour, suggesting an elevated tendency to form insoluble aggregates during the preparation of soy protein isolate. Small deformation rheological test revealed that the gelling times were shorter for those soy protein isolates derived from low lipoxygenase activity defatted soybean flours than that of high lipoxygenase activity. Frequency sweep showed that G′ of soy protein isolate derived from low lipoxygenase defatted soybean flour was independent of oscillation frequency in contrast to that of derived from non dry-heated defatted soybean flour, the latter showed a marked frequency dependence. Large deformation test revealed that the gel hardness increased about 10 times after dry heating of defatted soybean flour for 20 min. As the increase of the lipoxygenase residual activity, the gel permeability increased markedly, suggesting that soy protein isolate from high lipoxygenase defatted soybean flour produced coarser textured gel, which corresponded well with the results of scanning electron microscopy.
The effects of protein concentration, ionic strength, and lyophilization on heat-induced aggregation of soy proteins were analyzed by SDS-PAGE, SEC-HPLC and laser light scattering (LLS). SDS-PAGE profile suggested that the aggregates were formed via non-covalent forces and/or disulfide bonds. At ionic strength of zero, SEC-HPLC revealed that the samples were composed of three major fractions: aggregates, intermediate fractions and non-aggregated molecules. Furthermore, the relative proportion of the aggregate fraction increased as protein concentration increased. Similarly, LLS indicated that the average hydrodynamic radius (Rh) increased at higher protein concentration. In sample with an ionic strength of zero, the intermediate fraction decreased after freeze-drying with a concomitant increase of the aggregate fraction. When the sample was heated at elevated ionic strength, the SEC-HPLC and LLS profiles changed substantially, the intermediate fractions decreased, and lyophilization had effect on the fraction of aggregates. These experiments suggest novel strategies for producing soy protein aggregates with controlled properties.
... The unknown proteins in the soaking solutions were identied by mass spectrometry (MALDI-TOF/TOF-MS). 20 Briey, the protein spots on the Tricine-SDS-PAGE gel were excised and destained using 30% (v/v) acetonitrile containing 100 mM NH 4 HCO 3 . The destained gel was then washed with 100% acetonitrile and incubated in 10 mL reducing solution (100 mM DTT and 90 mL of 100 mM NH 4 HCO 3 ) for 30 min at 56 C. Aer washing with 100% acetonitrile, 70 mL of 100 mM NH 4 HCO 3 and 30 mL of 200 mM iodoacetic acid were added, and the mixture was incubated for 20 min in the dark. ...
... The adsorption of ex- ogenous proteins has Significant influence on the stability of OBs. It has recently been reported that the infestation of exogenous proteins may reduce the thermal stability of the oil body and promote the coalescence of the OB 26,27 . The amino acid contents of the OB proteins extracted from five peanut varieties are shown in Table 3. ...
... Mixtures of k-carrageenan and soy proteins under certain ionic conditions and polymer proportions may result on complex formation and synergistic effects between both biopolymers, ensuing improved textures and gel viscoelasticities, with both polymers contributing to the network formation (Baeza, Carp, Pérez, & Pilosof, 2002;Ortiz, Puppo, & Wagner, 2004). Nevertheless, when both biopolymers carry a negative charge, mixtures of k-carrageenan and soy proteins (both native protein and heat-induced aggregates) showed segregative phase separation (Li, Hua, Qiu, Yang, & Cui, 2008). ...
... However, few data have been reported on the fractionation of soybean glycinin isolated from LRSF. In previous papers [10,11], we reported that when defatted soybean flour with LOX activity were submitted to water extraction under mildly alkaline conditions in the preparation of soybean protein isolate, LOX may catalyze the oxidation of residual lipids present in soybean flour to initiate free-radical chain aggregation, causing undesirable changes in the nutritional and functional properties of the proteins. As a part of the above research, we present in this paper the effect of processing conditions on yields and purity of glycinin (11S) isolated from LRSF. ...
... This indicated that the various rheological properties of gel aggregates were reflection of the fractal structure of gel aggregate [34,38,46,60,68]. The salt-induced cold gelation process generally resulted in weak-link gels [38,46,51], whereas acid-induced cold gelation generally produced strong-link gel when the initial pH of protein solution was above 7.0 and weak-link gel when pH is below 7.0 [8,32,34,51,90,95]. The strong-link gels were generally transparent (fine- stranded; gel formed at low ionic strength and pH far from pI), while weak-link gels were turbid (particulate; gel formed at high ionic strength and pH close to pI). ...
... Walnuts are mostly consumed as snacks and are used in food formulations as an ingredient. Walnut milk beverage has been developed ( Chen et al. 2014). Walnut slurry obtained from wet grinding may be used as an ingredient to develop dairy products enriched with PUFAs. ...
... Wang & Damo- daran (1990) found that disulfide linked polymers played an indispensable role in gel formation of glycinin or soy protein isolate gels. Hua et al. (2005) reported an opposite result that for all soy protein isolates examined, the gel strength increased when Nethylmaleimide (NEM) was used to block sulfhydryl groups in protein, indicating that the different gelling behaviour of soy protein isolates would be attributed to their differences in surface hydrophobicity. In whey protein gel systems, Nguyen et al. (2014) found that gels made by NEM blocked protein had higher firmness, but these gels could be ruptured more easily than those with more disulfide bonds. ...
... The adsorption of ex- ogenous proteins has Significant influence on the stability of OBs. It has recently been reported that the infestation of exogenous proteins may reduce the thermal stability of the oil body and promote the coalescence of the OB 26,27 . The amino acid contents of the OB proteins extracted from five peanut varieties are shown in Table 3. ...
... The oil bodies contained only ∼50 g of lipids for 100 g which is in agreement with a previous study (White et al., 2006). Compared with the oat oils, oil bodies contained lower amount of phytosterols (∼230 and 75 mg, for the oils and oil bodies, respectively), suggesting that the phytosterols are present mainly in the oil phase although a small amount may be embedded within the monolayers of phospholipids and proteins (oleosin) (Chen, Cao, Zhao, Kong, & Hua, 2014). A significant proportion of the phytosterols present in the oils could have originated from the cell membranes when extracting the oil from the oat tissue (Hartmann, 1998). ...
Reference: Oat and lipolysis: Food matrix effect
... However, by increasing the temperature (>80 • C), the concentration of free SH groups decreased significantly. The decreased free SH content for SPI-Ex90 and SPI-Ex100 suggested that some cysteine could be lost because of SH oxidation, forming sulfur oxidation products other than disulfide bonds . These sulfur oxidation products are barely compressed by 2-mercaptoethanol in the subsequent determination of SH contents. ...